Alpha-aryl-N-alkylnitrones and pharmaceutical compositions containing the same

ABSTRACT

Disclosed are novel α-aryl-N-alkylnitrone compounds and pharmaceutical compositions containing such compounds. The disclosed compositions are useful as therapeutics for preventing and/or treating neurodegenerative, autoimmune and inflammatory conditions in mammals and as analytical reagents for detecting free radicals.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application Ser.No. 60/062,324, filed Oct. 17, 1997; U.S. Provisional Application Ser.No. 60/063,736, filed Oct. 29, 1997; and U.S. Provisional ApplicationSer. No. 60/090,475, filed Jun. 24, 1998. These applications areincorporated herein by reference in their entirety.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention relates to novel α-aryl-N-alkylnitrones and their use astherapeutic agents and analytical reagents. More particularly, thisinvention concerns novel α-aryl-N-alkylnitrones and their use astherapeutics for treating and/or preventing neurological, autoimmune andinflammatory conditions in mammals and as analytical reagents fordetecting free radicals.

2. State of the Art

Alzheimer's disease is a neurodegenerative condition in which nervecells in the brain are systematically destroyed resulting in progressivememory loss, mental confusion and ultimately death. The NationalInstitute on Aging (NIA) has recently estimated that about 4 millionpeople in the United States are currently afflicted with Alzheimer'sdisease. At present, there is no treatment that effectively prevents thedisease or reverses its symptoms.

In recent years, significant progress has been made in understanding thepathogenesis of Alzheimer's disease. For example, it is now known thatpatients with Alzheimer's disease develop amyloid plaque deposits aroundand between the nerve cells of their brain. These plaque deposits aremade up of fibrillar aggregates of a small peptide called amyloidβ-peptide or Aβ. The plaque deposits initially form in the hippocampusand cortical regions of the brain (areas associated with memory andcognition) and then spread to other areas as the disease progresses. Thedeposition of fibrils and plaques is also followed by inflammation ofthe surrounding support cells, called glia, which may lead to furtherloss of neurons. Eventually, the nerve cells in the brains of mostAlzheimer's patients develop tangles of a microtubule-associatedprotein, called tau, which are believed to be a response by the nervecells to damage.

Progress in understanding the underlying mechanisms of Alzheimer'sdisease has led to the development of various in vitro and in vivomodels to identify compounds effective for preventing and/or treatingAlzheimer's disease and other neurodegenerative conditions. In one suchin vitro model, compounds are evaluated for their ability to intervenein Aβ(140) or Aβ(1-42) beta-pleated sheet formation. Since thedeposition of amyloid β-peptide is associated with the development ofAlzheimer's disease, compounds which effectively disrupt the formationof Aβ(140) beta-pleated sheets are potentially useful for preventingand/or reversing Alzheimer's disease-related amyloid deposits.

In another in vitro model, compounds are evaluated for their ability toprotect against Aβ(25-35)-induced neuronal cell loss in rat embryonichippocampal neuronal/astrocyte cultures. As discussed above, patientswith. Alzheimer's disease suffer a progressive loss of neuronal cells.Accordingly, compounds which are effective in this in vitro test arepotentially useful for reducing or preventing neuronal cell loss inpatients afflicted with Alzheimer's disease or other neurodegenerativeconditions.

A third in vitro Alzheimer's disease model is based on the observationthat β-amyloid increases the release of cytokines, such asinterleukin-1β (IL-162 ), interleukin-6 (IL-6) and tumor necrosisfactor-α (TNFα), in human monocyte cells induced with lipopolysaccharide(LPS). IL-1β, IL-6 and TNFα are proteins associated with inflammatoryand immune responses. As previously mentioned, the deposition of fibrilsin the brains of Alzheimer's patients is associated with inflammation ofthe surrounding support cells. See, S. D. Yan et al., Proc. Natl. Acad.Sci. USA, 94, 5296 (1997). Thus, compounds effective in this in vitrotest are potentially useful for reducing and/or preventing theinflammation associated with Alzheimer's disease.

Additionally, elevated levels of IL-1β, IL-6, TNFα and other cytokinesare associated with a wide variety of inflammatory and autoimmuneconditions, including septic shock, rheumatoid arthritis, erythemanodosum leprosy, meningococcal meningitis, multiple sclerosis, systemiclupus and the like. See, L. Sekut et al., Drug News Perspect. 1196, 9,261; and A. Waage et al., J. Exp. Med. 1989, 170, 1859-1867.Accordingly, compounds which inhibit the production of such cytokinesare potentially useful for treating such inflammatory and autoimmuneconditions.

Similarly, various in vivo disease models are available for identifyingcompounds useful for preventing and/or treating neurodegenerative,autoimmune and inflammatory conditions. One such in vivo disease modelis based on the observation that mammals suffer cognitive impairmentwhen Aβ(25-35) and ibotenate are injected into the hippocampus of theirbrain. Since amyloid β-peptide deposits are associated with Alzheimer'sdisease, compounds which effectively reduce the cognitive impairmentcaused by Aβ(25-35)/ibotenate are potentially useful for the preventionand/or treatment of Alzheimer's disease and other neurodegenerativeconditions. Another in vivo disease model is based on the observationthat certain strains of autoimmune mice develop cognitive deficits asthey mature. See, for example, Forster et al., Behav. Neural Biology1988, 49, 139-151. Thus, compounds which prevent or reduce suchcognitive deficits are potentially useful for preventing and/or treatingneurodegenerative and autoimmune conditions.

It has now been discovered that certain novel α-aryl-N-alkylnitronecompounds effectively inhibit thee formation of Aβ(142) beta-pleatedsheets and/or protect against neuronal cell loss and/or inhibit therelease of cytokines, such as IL-1β and TNFα. Additionally, in in vivotests, these compounds have been found to reduce the cognitiveimpairment caused by Aβ(25-35)/ibotenate and to reduce the cognitivedeficits that develop in certain strains of autoimmune mice.Accordingly, such compounds are useful for the prevention and/ortreatment of neurodegenerative, autoimmune and inflammatory conditionsin mammals.

The α-aryl-N-alkylnitrone compounds of this invention are also useful asanalytical reagents for detecting free radicals. In this regard, thecompounds of this invention function as “spin traps” by reacting withunstable free radicals to form relatively stable free radical spinadducts which are observable by, electron spin resonance (ESR)spectroscopy. Accordingly, when used as spin traps, the compounds ofthis invention allow free radicals to be identified and studied usingESR and related techniques.

SUMMARY OF THE INVENTION

This invention provides novel α aryl-N-alkylnitrone compounds which areuseful as therapeutics for treating and/or preventing neurological,autoimmune and inflammatory conditions in mammals and as analyticalreagents for detecting free radicals. In particular, the compounds ofthis invention are useful for preventing and/or treating Alzheimer'sdisease.

Accordingly, in one of its composition aspects, this invention isdirected to compounds of formula I:

wherein

-   -   R¹ is selected from the group consisting of alkoxy, alkaryloxy,        alkcycloalkoxy, aryloxy, and cycloalkoxy;    -   R² is selected from the group consisting of hydrogen, alkoxy,        alkcycloalkoxy, cycloalkoxy and halogen, or when R¹ and R² are        attached to adjacent carbon atoms, R¹ and R² may be joined        together to form an alkylenedioxy group;    -   R³ is selected from the group consisting of hydrogen, alkoxy,        alkcycloalkoxy, cycloalkoxy and halogen;    -   R⁴ is selected from the group consisting of hydrogen and alkyl;    -   R⁵ is selected from the group consisting of alkyl having at        least 3 carbon atoms, substituted alkyl having at least 3 carbon        atoms and cycloalkyl;    -   provided that:    -   (i) when R² and R³ are independently hydrogen or methoxy, R¹ is        not methoxy;    -   (ii) when R², R³ and R⁴ are hydrogen and R⁵ is tert-butyl, then        R¹ is not 4-n-butoxy, 4-n-pentyloxy or 4-n-hexyloxy;    -   (iii) when R², R³ and R⁴ are hydrogen and R⁵ is isopropyl, then        R¹ is not 4-ethoxy;    -   (iv) when R¹ and R² are joined together to form a        3,4-methylenedioxy group and R³ and R⁴ are hydrogen, then R⁵ is        not isopropyl or tert-butyl;    -   (v) when R², R³ and R⁴ are hydrogen and R⁵ is        1-hydroxy-2-methylprop-2-yl, then R¹ is not 2-ethoxy;    -   (vi) when R¹ is 4-methoxy, R² is 3-ethoxy, and R³ and R⁴ are        hydrogen, then R⁵ is not 2,2-dimethylbut-3-yl or        1-hydroxy-2-methylprop-2-yl; and    -   (vii) when R³ and R⁴ are hydrogen and R⁵ is tert-butyl, then R¹        is not 4-methoxy when R² is 2-fluoro, and R¹ is not 2-methoxy        when R² is 4-fluoro.

Preferably, in the compounds of formula I above, R¹ is selected from thegroup consisting of alkoxy, alkaryloxy and cycloalkoxy. More preferably,R¹ is alkoxy having 1 to about 8 carbon atoms or alkaryloxy having 7 toabout 10 carbon atoms. Particularly preferred R¹ groups include methoxy,ethoxy, butoxy, pentyloxy, hexyloxy, cyclohexyloxy, heptyloxy, octyloxy,benzyloxy, 4-fluorobenzyloxy and 4-methoxybenzyloxy.

R² is preferably selected from the group consisting of hydrogen, alkoxyand fluoro. More preferably, R² is hydrogen, alkoxy having 2 to about 8carbon atoms, or fluoro. Particularly preferred R² groups includehydrogen, ethoxy and fluoro.

When R¹ and R² are attached to adjacent carbon atoms, R¹ and R² are alsopreferably joined together to form an alkylenedioxy group having 1 toabout 6 carbon atoms. Particularly preferred alkylenedioxy groupsinclude methylenedioxy and ethylenedioxy, provided that when R¹ and R²are joined together to form a 3,4-methylenedioxy group and R³ and R⁴ arehydrogen, then R⁵ is not isopropyl or tert-butyl.

Preferably, R³ is hydrogen or alkoxy. More preferably, R³ is hydrogen oralkoxy having 2 to 8 carbon atoms. Particularly preferred R groupsinclude hydrogen and ethoxy.

R⁴ is preferably hydrogen or lower alkyl. More preferably, R⁴ ishydrogen or alkyl having 1 to 4 carbon atoms. Still more preferably, R⁴is hydrogen.

R⁵ is-preferably selected from the group consisting of alkyl having 3 toabout 8 carbon atoms, substituted alkyl having 3 to 8 carbon atoms andcycloalkyl having 3 to about 10 carbon atoms. More preferably, R⁵ isalkyl having 3 to 6 carbon atoms or cycloalkyl having 5 to 6 carbonatoms.

Particularly preferred R⁵ groups include n-propyl, isopropyl,1-methoxy-2-methylproo-2-yl, n-butyl, but-2-yl, tert-butyl,2-methylbut-2-yl, 3-methylbut-1-yl, 3,3-dimethylbut-2-yl,4-methylpent-2-yl, 2,4-dimethyl-2-pentyl, 2,2,4,4-tetramethylpent-3-yl,cyclopropyl, cyclobutyl, tert-octyl (2,4,4-trimethylpent-2-yl),cyclopentyl, cyclohexyl, cyclooctyl, 1-adamantyl, 2-adamantyl,3,5-dimethyl-l-adamantyl and benzyl. When R⁵ is adamantyl, 1-adamantylis preferred.

Other suitable R⁵ groups include, by way of example, 1-phenylethyl,1-phenylprop-2-yl, 2-phenylprop-2-yl, 2-benzylprop-2-yl,2-(methoxycarbonyl)-prop-2-yl, 1,3-dihydroxy-2-(hydroxymethyl)prop-2-yl,1-sulfo-2-methylprop-2-yl, 4-fluorobenzyl, 3,4-dimethoxybenzyl,3-thiomethoxybut-1-yl and 3-thiomethoxyprop-1-yl.

An especially preferred group of compounds of formula I are those inwhich R¹ is a 2-ethoxy group; R², R³ and R⁴ are each hydrogen; and R⁵ isas defined above.

Another especially preferred group of compounds of formula I are thosein which R¹ is a 4-ethoxy group; R², R³ and R⁴ are each hydrogen; and R⁵is as defined above.

Still another especially preferred group of compounds of formula I arethose in which R¹ is a 4-benzyloxy group; R², R³ and R⁴ are eachhydrogen; and R⁵ is as defined above.

Yet another especially preferred group of compounds of formula I arethose in which R¹ is a 3-ethoxy group; R² is a 4-methoxy group; R³ andR⁴ are each hydrogen; and R⁵ is as defined above.

In a preferred embodiment, this invention is directed to a compound offormula II:

wherein

-   -   R⁶ is selected from the group consisting of alkoxy having 1 to 8        carbon atoms, alkaryloxy having 7 to 10 carbon atoms, aryloxy        having 6 to 10 carbon atoms and cycloalkoxy having 3 to 10        carbon atoms;    -   R⁷ is selected from the group consisting of alkoxy having 1 to 8        carbon atoms and fluoro, or when R⁶ and R⁷ are attached to        adjacent carbon atoms, R⁶ and R⁷ may be joined together to form        an alkylenedioxy group having 1 to about 6 carbon atoms;    -   R⁸ is selected from the group consisting of hydrogen and alkoxy        having 1 to 8 carbon atoms; and    -   R⁹ is selected from the group consisting of alkyl having 3 to        about 8 carbon atoms, substituted alkyl having 3 to about 8        carbon atoms and cycloalkyl having 3 to about 10 carbon atoms;    -   provided that:    -   (i) when R⁷ is methoxy and R⁸ is hydrogen or methoxy, R⁶ is not        methoxy;    -   (ii) when R⁶ and R⁷ are joined together to form a        3,4-methylenedioxy group and R⁸ is hydrogen, then R⁹ is not        isopropyl or tert-butyl; and    -   (iii) when R⁶ is 4-methoxy, R⁷ is 3-ethoxy and R⁸ is hydrogen,        then R⁹ is not 2,2-dimethylbut-3-yl or        1-hydroxy-2-methylprop-2-yl.

In a preferred embodiment, R⁶ is alkoxy having 1 to 8 carbon atoms, R⁷is alkoxy, having 2 to 8 carbon atoms and R⁸ is hydrogen. In thisembodiment, particularly preferred R⁶ groups include methoxy, ethoxy,butoxy, pentyloxy, hexyloxy, heptyloxy and octyloxy, and particularlypreferred R⁷ groups include ethoxy. More preferably, R⁶ is methoxy andR⁷ is ethoxy.

In another preferred embodiment, R⁶ is ethoxy; and R⁷ and R⁸ arehydrogen.

In yet another preferred embodiment, R⁶ is benzyloxy, R⁷ is alkoxyhaving 1 to 8 carbon atoms, and R⁸ is hydrogen. In this embodiment,particularly preferred R⁷ groups include methoxy, ethoxy, butoxy,pentyloxy, hexyloxy, heptyloxy and octyloxy. In another preferredembodiment, R⁶ is benzyloxy; and R⁷ and R⁸ are hydrogen.

In still another preferred embodiment, R⁶ is alkoxy having 1 to 8 carbonatoms, R⁷ is fluoro and R⁸ is hydrogen. In this embodiment, particularlypreferred R⁶ groups include methoxy, ethoxy, butoxy, pentyloxy,hexyloxy, heptyloxy and octyloxy.

In yet another preferred embodiment, R⁶ and R⁷ are joined together toform a methylenedioxy or ethylenedioxy group and R⁸ is hydrogen,provided that when R⁶ and R⁷ are joined together to form a3,4-methylenedioxy group and R⁸ is hydrogen, then R⁹ is not isopropyl ortert-butyl.

In the above embodiments, R⁹ is preferably alkyl having 3 to 6 carbonatoms or cycloalkyl having 5 to 10 carbon atoms. Particularly preferredR⁹ groups include n-propyl, isopropyl, 1-methoxy-2-methylproo-2-yl,n-butyl, but-2-yl, tert-butyl, 2-methylbut-2-yl, 3-methylbut-1-yl,3,3-dimethylbut-2-yl, 4-methylpent-2-yl, 2,4-dimethyl-2-pentyl,2,2,4,4-tetramethylpent-3-yl, cyclopropyl, cyclobutyl, tert-octyl(2,4,4-trimethylpent-2-yl), cyclopentyl, cyclohexyl, dyclooctyl,1-adamantyl, 2-adamantyl, 3,5-dimethyl-1-adamantyl, benzyl. When R⁹ isadamantyl, 1-admantyl is preferred. Especially-preferred R⁹ groups areisopropyl, tert-butyl, 2,4-dimethyl-2-pentyl, tert-octyl, 1-adamantyl,cyclopropyl and cyclohexyl.

In another of its composition aspects, this invention is directed toeach of the individual compounds:

-   -   α-(4-heptyloxyphenyl)-N-tert-butylnitrone    -   α-(4-hexyloxyphenyl)-N-n-propylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-tert-butylnitrone    -   α-(4-ethoxyphenyl)-N-tert-butylnitrone    -   α-(4-benzyloxy-3-methoxyphenyl)-N-tert-butylnitrone    -   α-[3-(4-methoxyphenoxy)phenyl]-N-tert-butylnitrone    -   α-(2-ethoxyphenyl)-N-tert-butylnitrone    -   α-(3,4-ethylenedioxyphenyl)-N-tert-butylnitrone    -   α-(4-ethoxyphenyl)-N-cyclohexylnitrone    -   α-(4-benzyloxy-3-methoxyphenyl)-N-cyclohexylnitrone    -   α-3-ethoxy-4-methoxyphenyl)-N-cyclohexylnitrone    -   α-(3,4-ethylenedioxyphenyl)-N-cyclohexylnitrone    -   α-4-ethoxy-3-methoxyphenyl)-N-cyclohexylnitrone    -   α-(3,4-ethylenedioxyphenyl)-N-isopropylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-isopropylnitrone    -   α-(2- ethoxyphenyl)-N-isopropylnitrone    -   α-(2-ethoxyphenyl)-N-cyclohexylnitrone    -   α-(4-benzyloxy-3-methoxyphenyl)-N-isopropylnitrone    -   α-(4-ethoxy-3-methoxyphenyl)-N-isopropylnitrone    -   α-(3-ethoxy-4-hexyloxyphenyl)-N-cyclohexylnitrone    -   α-(4-benzyloxy-3-methoxyphenyl)-N-n-butylnitrone    -   α-(4-ethoxy-3-methoxyphenyl)-N-n-butylnitrone,    -   α-(2- ethoxyphenyl)-N-n-butylnitrone    -   α-(3-ethoxy-α-methoxyphenyl)-N-n-butylnitrone    -   α-(3-ethoxy-4-hexyloxyphenyl)-isopropylnitrone    -   α-(3-ethoxy-4-hexyloxyphenyl)-N-tert-butylnitrone    -   α-(2-fluoro-4-octyloxyphenyl)-N-tert-butylnitrone    -   α-(2,4,6-triethoxyphenyl)-N-tert-butylnitrone    -   α-(2,4,6-triethoxyphenyl)-N-cyclohexylnitrone    -   α-(2-n-butoxyphenyl)-N-tert-butylnitrone    -   α-(3,4-diethoxyphenyl)-N-tert-butylnitrone    -   α-(2-fluoro-4-heptyloxyphenyl)-N-tert-butylnitrone    -   α-(2-fluoro-4-ethoxyphenyl)-N-tert-butylnitrone    -   α-(2-fluoro-4-ethoxyphenyl)-N-cyclohexylnitrone    -   α-(2-ethoxyphenyl)-N-1-adamantylvitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-1-adamantylnitrone    -   α-(4-ethoxyphenyl)-N-cyclopentylnitrone    -   α-(4-ethoxyphenyl)-N-tert-octylnitrone    -   α-(4-benzyloxyphenyl)-N-tert-butylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclopentylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclohexylnitrone    -   α-(2-ethoxyphenyl)-N-cyclopentylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-tert-octylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(2,4-dimethyl-2-pentyl)nitrone    -   α-(4-ethoxyphenyl)-N-n-butylnitrone    -   α-(2-ethoxyphenyl)-N-benzylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(2,2,4,4-tetramethylpent-3-yl)nitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(4-methylpent-2-yl)nitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-but-2-ylnitrone    -   α-(2-ethoxyphenyl)-N-but-2-ylnitrone    -   α-[4-(4-fluorobenzyloxy)phenyl]-N-tert-butylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-cyclopentylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-n-propylnitrone    -   α-(4-benzyloxyphenyl)-N-n-propylnitrone    -   α-(4-benzyloxyphenyl)-N-isopropylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(2-methylbut-2-yl)nitrone    -   α-(2-ethoxyphenyl)-N-(2-methylbut-2-yl)nitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-cyclooctylnitrone    -   α-(2-ethoxyphenyl)-N-cyclobutylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-cyclobutylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclobutylnitrone    -   α-(4-benzyloxyphenyl)-N-tert-octylnitrone    -   α-[4-(4-fluorobenzyloxy)phenyl]-N-cyclohexylnitrone    -   α-(2-ethoxyphenyl)-N-tert-octylnitrone    -   α-[4-(4-fluorobenzyloxy)phenyl]-N-isopropylnitrone    -   α-(2-ethoxyphenyl)-N-cyclooctylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclopropylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-cyclopropylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclooctylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(3,5-dimethyl-1-adamantyl)nitrone    -   α-(4-benzyloxyphenyl)-N-1-adamantylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(1-methoxy-2-methylprop-2-yl)nitrone    -   α-(4-benzyloxyphenyl)-N-2-adamantylnitrone    -   α-(4-ethoxyphenyl)-N-cyclooctylnitrone    -   α-(4-ethoxyphenyl)-N-1-adamantylnitrone    -   α-[4-(4-methoxybenzyloxy)phenyl]-N-tert-butylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-(3-methylbut-1-yl)nitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-cyclooctylnitrone, and    -   α-[4-(4-fluorobenzyloxy)phenyl]-N-cyclopentylnitrone.

Particularly preferred compounds include:

-   -   α-(2-ethoxyphenyl)-N-tert-butylnitrone    -   α-(2-ethoxyphenyl)-N-cyclohexylnitrone    -   α-4-ethoxyphenyl)-N-cyclohexylnitrone    -   α-(4-benzyloxyphenyl)-N-tert-butylnitrone    -   α-(4-benzyloxyphenyl)-N-cyclopentylnitrone    -   α-(3-ethoxy-4-methoxyphenyl)-N-adamantylnitrone, and    -   α-(3-ethoxy-4-methoxyphenyl)-N-tert-octylnitrone.

In another of its composition aspects, this invention is directed topharmaceutical compositions comprising a pharmaceutically acceptablecarrier and a pharmaceutically effective amount of a compound of formulaI:

wherein R¹-R⁵ are as defined above.

In additional composition aspects, this invention is directed topharmaceutical compositions comprising a pharmaceutically acceptablecarrier and a pharmaceutically effective amount of a compound of formulaII above.

As previously mentioned, the α-aryl-N-alkylnitrone compounds of thisinvention have been discovered to inhibit the formation of Aβ(142)beta-pleated sheets and/or to protect against Aβ(25-35)-induced neuronalcell loss and/or to reduce β-amyloid-induced release of cytokines, suchas IL-1β and TNFα, in human monocyte cells. Such compounds have alsobeen found to reduce the cognitive defects caused by Aβ(25-35)/ibotenateas well as those which develop in certain strains of autoimmune mice.Compounds having such properties are useful for preventing and/ortreating neurodegenerative, autoimmune and inflammatory conditions.

Accordingly, in one of its method aspects, this invention is directed toa method for treating a patient with a neurodegenerative disease whichmethod comprises administering to said patient a pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier and aneffective neurodegenerative disease-treating amount of a compound offormula I or formula II above.

In another of its method aspects, this invention is directed to a methodfor preventing the onset of a neurodegenerative disease in a patient atrisk for developing the neurodegenerative disease which method comprisesadministering to said patient a pharmaceutical composition comprising apharmaceutically acceptable carrier and an effective neurodegenerativedisease-preventing amount of a compound of formula I or formula IIabove.

In preferred embodiments of this invention, the neurodegenerativedisease treated and/or prevented in Lie above methods is Alzheimer'sdisease, Parkinson's disease, HIV dementia and the like.

In still another of its method aspects, this invention is directed to amethod for treating a patient with an autoimmune disease which methodcomprises administering to said patient a pharmaceutical compositioncomprising a pharmaceutically acceptable carrier and an effectiveautoimmune disease-treating amount of a compound of formula I or formulaII above.

In yet another of its method aspects, this invention is directed to amethod for preventing the onset of an autoimmune disease in a patient atrisk for developing the autoimmune disease which method comprisesadministering to said patient a pharmaceutical composition comprising apharmaceutically acceptable carrier and an effective autoimmunedisease-preventing amount of a compound of formula I or formula IIabove.

In preferred embodiments of this invention, the autoimmune diseasetreated and/or prevented in the above methods is systemic lupus,multiple sclerosis and the like.

In still another of its method aspects, this invention is directed to amethod for treating a patient with an inflammatory disease which methodcomprises administering to said patient a pharmaceutical composition acomprising a pharmaceutically acceptable carrier and an effectiveinflammatory disease-treating amount of a compound of formula I orformula II above.

In yet another of its method aspects, this invention is directed to amethod for preventing the onset of an inflammatory disease in a patientat risk for developing the inflammatory disease which method comprisesadministering to, said patient a pharmaceutical composition comprising apharmaceutically acceptable carrier and an effective inflammatorydisease-preventing amount of a compound of formula I or formula IIabove.

In preferred embodiments of this invention, the inflammatory diseasetreated and/or prevented in the above methods is rheumatoid arthritis,septic shock, erythema nodosum leprosy, septicemia, adult respiratorydistress syndrome (ARDS), inflammatory bowel disease (IBD), uveitis andthe like.

In another of its aspects, this invention is directed to the use of acompound of formula I or formula II above in the manufacture of aformulation or medicament for a medicinal treatment. Preferably, themedical treatment is the therapeutic or prophylactic treatment of aneurodegenerative disease, an autoimmune disease or an inflammatorydisease.

Particularly preferred compounds include those represented in Tables Iand II below. TABLE I

R^(a) R^(b) R^(c) R^(d) H— H— CH₃(CH₂)₆—O— (CH₃)₃C— H— H— CH₃(CH₂)₅—O—CH₃CH₂CH₂— H— CH₃CH₂—O— CH₃—O— (CH₃)₃C— H— H— CH₃CH₂—O— (CH₃)₃C— H—CH₃—O— PhCH₂—O— (CH₃)₃C— H— 4-(CH₃—O)-Ph-O— H— (CH₃)₃C— CH₃CH₂—O— H— H—(CH₃)₃C— H— —O—CH₂CH₂—O— (CH₃)₃C— H— H— CH₃CH₂—O— cyclohexyl- H— CH₃—O—PhCH₂—O— cyclohexyl- H— CH₃CH₂—O— CH₃—O— cyclohexyl- H— —O—CH₂CH₂—O—cyclohexyl- H— CH₃—O— CH₃CH₂—O— cyclohexyl- H— —O—CH₂CH₂—O— (CH₃)₂CH— H—CH₃CH₂—O— CH₃—O— (CH₃)₂CH— CH₃CH₂—O— H— H— (CH₃)₂CH— CH₃CH₂—O— H— H—cyclohexyl- H— CH₃—O— PhCH₂—O— (CH₃)₂CH— H— CH₃—O— CH₃CH₂—O— (CH₃)₂CH—H— CH₃CH₂—O— CH₃(CH₂)₅—O— cyclohexyl- H— CH₃—O— PhCH₂—O— CH₃(CH₂)₃— H—CH₃—O— CH₃CH₂—O— CH₃CH₃(CH₂)₃— CH₃CH₂—O— H— H— CH₃(CH₂)₃— H— CH₃CH₂—O—CH₃—O— CH₃(CH₂)₃— H— CH₃CH₂—O— CH₃(CH₂)₅—O— (CH₃)₂CH— H— CH₃CH₂—O—CH₃(CH₂)₅—O— (CH₃)₃C— F— H— CH₃(CH₂)₇—O— (CH₃)₃C— CH₃(CH₂)₃—O— H— H—(CH₃)₃C— H— CH₃CH₂—O— CH₃CH₂—O— (CH₃)₃C— F— H— CH₃(CH₂)₅—O— (CH₃)₃C— F—H— CH₃CH₂—O— (CH₃)₃C— F— H— CH₃CH₂—O— cyclohexyl- CH₃CH₂—O— H— H—1-adamantyl- H— CH₃CH₂—O— CH₃—O— 1-adamantyl- H— H— CH₃CH₂—O—cyclopentyl- H— H— CH₃CH₂—O— (CH₃)₃CCH₂— (CH₃)₂C— H— H— PhCH₂—O—(CH₃)₃C— H— H— PhCH₂—O— cyclopentyl- H— H— PhCH₂—O— cyclohexyl-CH₃CH₂—O— H— H— cyclopentyl- H— CH₃CH₂—O— CH₃—O— (CH₃)₃CCH₂— (CH₃)₂C— H—CH₃CH₂—O— CH₃—O— (CH₃)₂CHCH₂— (CH₃)₂C— H— H— CH₃CH₂—O— CH₃(CH₂)₃—CH₃CH₂—O— H— H— PhCH₂— H— CH₃CH₂—O— CH₃—O— [(CH₃)₃C]₂CH— H— CH₃CH₂—O—CH₃—O— (CH₃)₂CHCH₂— (CH₃)CH— H— CH₃CH₂—O— CH₃—O— CH₃CH₂(CH₃)— CH—CH₃CH₂—O— H— H— CH₃CH₂(CH₃)— CH— H— H— 4-F-PhCH₂—O— (CH₃)₃C— H—CH₃CH₂—O— CH₃—O— cyclopentyl- H— CH₃CH₂—O— CH₃—O— CH₃CH₂CH₂— H— H—PhCH₂—O— CH₃CH₂CH₂— H— H— PhCH₂—O— (CH₃)₂CH— H— CH₃CH₂—O— CH₃—O—CH₃CH₂(CH₃)₂C— CH₃CH₂—O— H— H— CH₃CH₂(CH₃)₂C— H— CH₃CH₂—O— CH₃—O—cyclooctyl- CH₃CH₂—O— H— H— cyclobutyl- H— CH₃CH₂—O— CH₃—O— cyclobutyl-H— H— PhCH₂—O— cyclobutyl- H— H— PhCH₂—O— (CH₃)₃CCH₂— (CH₃)₂C— H— H—4-F-PhCH₂—O— cyclohexyl— CH₃CH₂—O— H— H— (CH₃)₃CCH₂— (CH₃)₂C— H— H—4-F-PhCH₂—O— (CH₃)₂CH— CH₃CH₂—O— H— H— cyclooctyl- H— H— PhCH₂—O—cyclopropyl- H— CH₃CH₂—O— CH₃—O— 2-adamantyl- H— CH₃CH₂—O— CH₃—O—cyclopropyl- H— H— PhCH₂—O— cyclooctyl- H— CH₃CH₂—O— CH₃—O—3,5-di(CH₃)-1- adamantyl H— H— PhCH₂—O— 1-adamantyl- H— CH₃CH₂—O— CH₃—O—CH₃OCH₂— (CH₃)₂C— H— H— PhCH₂—O— 2-adamantyl- H— H— CH₃CH₂—O—cyclooctyl- H— H— CH₃CH₂—O— 1-adamantyl- H— H— 4-CH₃O-PhCH₂O— (CH₃)₃C—H— CH₃CH₂—O— CH₃—O— (CH₃)₂CHCH₂— CH₂— H— CH₃CH₂—O— CH₃—O— cyclooctyl- H—H— 4-F-PhCH₂—O— cyclopentyl-

TABLE II

R^(e) R^(f) R^(g) R^(h) CH₃CH₂—O— CH₃CH₂—O— CH₃CH₂—O— (CH₃)₃C— CH₃CH₂—O—CH₃CH₂—O— CH₃CH₂—O— cyclohexyl-

BRIEF DESCRIPTION OF THE DRAWING

FIG. 1 is an electron spin resonance (ESR) spectra of the radical adductof α-(2-ethoxyphenyl)-N-tert-butylnitrone and a methyl radical.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

When describing the α-aryl-N-alkylnitrones, pharmaceutical compositionsand methods of this invention, the following terms have the followingmeanings:

The term “β-amyloid peptide” refers to a 39-43 amino acid peptide havinga molecular weight of about 4.2 kD, which peptide is substantiallyhomologous to the form of the protein described by Glenner, et al.,Biochem. Biophys. Res. Commun., 120:885-890. (1984), including mutationsand post-translational modifications of the normal β-amyloid peptide.

The term “cytokines” refers to peptide protein mediators that areproduced by immune cells to modulate cellular functions. Examples ofcytokines include, interleukin-1β (IL-1β), interleukin-6 (IL-6) andtumor necrosis factor-α (TNFα).

“Acyl” refers to the group —OC(O)R where R is alkyl or aryl.

“Alkyl” refers to monovalent alkyl groups preferably having from 1 toabout 10 carbon atoms, more preferably 1 to 8 carbon atoms and stillmore preferably 1 to 6 carbon atoms. This term is exemplified by groupssuch as methyl, ethyl, n-propyl, isopropyl, n-butyl; isobutyl,tert-butyl, n-hexyl, n-octyl, tert-octyl and the like. The term “loweralkyl” refers to alkyl groups having 1 to 6 carbon atoms.

“Substituted alkyl” refers to an alkyl group, preferably of from 1 to 10carbon atoms, having from 1 to 5 substituents, and preferably 1 to 3substituents, selected from the group consisting of alkoxy, cycloalkyl,cycloalkoxy, acyl, aminoacyl, amino, aminocarbonyl, cyano, halogen,hydroxyl, carboxyl, keto, thioketo, alkoxycarbonyl, thiol, thioalkoxy,aryl, aryloxy, nitro, —OSO₃H and pharmaceutically acceptable saltsthereof, —SO-alkyl, —SO-substituted alkyl, —SO-aryl, —SO₂-alkyl,—SO₂-substituted-alkyl, —SO₂-aryl, and mono- and di-alkylamino, mono-and di-arylamino, and unsymmetric di-substituted amines having differentsubstituents selected from alkyl, substituted alkyl and aryl.

“Alkylene” refers to divalent alkylene groups preferably having from 1to 10 carbon atoms and more preferably 1 to 6 carbon atoms which can bestraight chain or branched. This term is exemplified by groups such asmethylene (—CH₂—), ethylene (—CH₂CH₂—), the propylene isomers (e.g.,—CH₂CH₂CH₂— and —CH(CH₃)CH₂—) and the like.

“Alkylenedioxy” refers to —O-alkylene-O— groups preferably having from 1to 10 carbon atoms and more preferably 1 to 6 carbon atoms which can bestraight chain or branched. This term is exemplified by groups such asmethylenedioxy (—OCH₂O—), ethylenedioxy (—OCH₂CH₂O—) and the like.

“Alkenylene” refers to divalent alkenylene groups preferably having from2 to 10 carbon atoms and more preferably 2 to 6 carbon atoms which canbe straight chain or branched and having at least 1 and preferably from1-2 sites of alkenyl unsaturation. This term is exemplified by groupssuch as ethenylene (—CH═CH—), the propenylene isomers (e.g., —CH═CHCH₂—and —C(CH₃)═CH—and —CH═C(CH₃)—) and the like.

“Alkaryl” refers to -alkylene-aryl groups preferably having from 1 to 10carbon atoms in the alkylene moiety and from 6 to 14 carbon atoms in thearyl moiety. Such alkaryl groups are exemplified by benzyl, phenethyl,and the like.

“Alkaryloxy” refers to —O -alkylene-aryl groups preferably having from 1to 10 carbon atoms in the alkylene moiety and from 6 to 14 carbon atomsin the aryl moiety. Such alkaryl groups are exemplified by benzyloxy,4-fluorobenzyloxy, phenethyloxy, and the like.

“Alkcycloalkyl” refers to -alkylene-cycloalkyl groups preferably havingfrom 1 to 10 carbon atoms in the alkylene moiety and from 3 to 8 carbonatoms in the cycloalkyl moiety. Such alkcycloalkyl groups areexemplified by —CH₂-cyclopropyl, —CH₂-cyclopentyl, —CH₂CH₂-cyclohexyl,and the like.

“Alkcycloalkoxy” refers to —O-alkylene-cycloalkyl groups preferablyhaving from 1 to 10 carbon atoms in the alkylene moiety and from 3 to 8carbon atoms in the cycloalkyl moiety. Such alkcycloalkoxy groups areexemplified by —OCH₂-cyclopropyl, —OCH₂-cyclopentyl,—OCH₂CH₂-cyclohexyl, and the like.

“Alkoxy” refers to the group “alkyl-O—”. Preferred alkoxy groupsinclude, by way of example, methoxy, ethoxy, n-propoxy, isopropoxy,n-butoxy, tert-butoxy, sec-butoxy, n-pentyloxy, n-hexyloxy,1,2-dimethylbutoxy, and the like.

“Alkoxycarbonyl” refers to the group —C(O)OR where R is alkyl.

“Alkenyl” refers to alkenyl groups preferably having from 2 to 10 carbonatoms and more preferably 2 to 6 carbon atoms and having at least 1 andpreferably from 1-2 sites of alkenyl unsaturation. Preferred alkenylgroups include ethenyl (—CH═CH₂), n-propenyl (—CH₂CH═CH₂), isopropenyl(—C(CH₃)═CH₂), and the like.

“Alkynyl” refers to alkynyl groups preferably having from 2 to 10 carbonatoms and more preferably 2 to 6 carbon atoms and having at least 1 andpreferably from 1-2 sites of alkynyl unsaturation. Preferred alkynylgroups include ethynyl (—C≡—CH), propargyl (—CH₂C≡CH), and the like.

“Aminocarbonyl” refers to the group —C(O)NRR where each R isindependently hydrogen or alkyl.

“Aminoacyl” refers to the group —NRC(O)R where each R is independentlyhydrogen or alkyl.

“Aryl” refers to an unsaturated aromatic carbocyclic group of from 6 to14 carbon atoms having a single ring (e.g., phenyl) or multiplecondensed rings (e.g., naphthyl or anthryl). Preferred aryls includephenyl, naphthyl and the like. Unless otherwise constrained by thedefinition for the individual substituent, such aryl groups canoptionally be substituted with from 1 to 3 substituents selected fromthe group consisting of alkyl, alkoxy, alkaryloxy, alkenyl, alkynyl,amino, aminoacyl, aminocarbonyl, alkoxycarbonyl, aryl, carboxyl,cycloalkoxy, cyano, halo, hydroxy, nitro, trihalomethyl, thioalkoxy, andthe like.

“Aryloxy” refers to —O-aryl groups wherein “aryl” is as defined above.

“Carboxyl” refers to the group —C(O)OH.

“Cyano” refers to the group —CN.

“Cycloalkyl” refers to cyclic alkyl groups of from 3 to 10 carbon atomshaving a single cyclic ring or multiple condensed rings, including fusedand bridged ring systems, which can be optionally substituted with from1 to 3 alkyl groups. Such cycloalkyl groups include, by way of example,single ring structures such as cyclopropyl, cyclobutyl, cyclopentyl,cyclooctyl, 1-methylcyclopropyl, 2-methylcyclopentyl,2-methylcyclooctyl, and the like, or multiple ring structures such asadamantyl, and the like.

“Cycloatkoxy” refers to —O-cycloalkyl groups. Such cycloalkoxy groupsinclude, by way of example, cyclopentyloky, cyclohexyloxy and the like.

“Cycloalkenyl” refers to cyclic alkenyl groups of from 4 to 10 carbonatoms having a single cyclic ring and at least one point of internalunsaturation which can be optionally substituted with from 1 to 3 alkylgroups. Examples of suitable cycloalkenyl groups include, for instance,cyclopent-3-enyl, cyclohex-2-enyl, cyclooct-3-enyl and the like.

“Halo” or “halogen” refers to fluoro, chloro, bromo and iodo. Preferredhalo groups are either fluoro or chloro.

“Keto” or “oxo” refers to the group ═O.

“Nitro” refers to the group —NO₂.

“tert-Octyl” refers to a 2,4,4-trimethyl-2-pentyl group.

“Thiol” refers to the group —SH.

“Thioalkoxy” refers to the group —S-alkyl.

“Thioketo” refers to the group ═S.

“Pharmaceutically acceptable salt” refers to pharmaceutically acceptablesalts which are derived from a variety of organic and inorganiccounter-ions well known in the art and include, by way of example only,sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, andthe like; and when the molecule contains a basic functionality, salts oforganic or inorganic acids, such as hydrochloride, hydrobromide,tartrate, mesylate, acetate, maleate, oxalate and the like. The term“pharmaceutically acceptable cation” refers to a pharmaceuticallyacceptable cationic counter-ion of an acidic functional group. Suchcations are exemplified by sodium, potassium, calcium, magnesium,ammonium, tetraalkylammonium cations, and the like.

General Synthetic Procedures

The α-aryl-N-alkylnitrones of this invention can be prepared fromreadily available starting materials using the following general methodsand procedures. It will be appreciated that where typical or preferredprocess conditions (i.e., reaction temperatures, times, mole ratios ofreactants, solvents, pressures, etc.) are given, other processconditions can also be used unless otherwise stated. Optimum reactionconditions may vary with the particular reactants or solvent used, butsuch conditions can be determined by one skilled in the art by routineoptimization procedures.

Additionally, as will be apparent to those skilled in the art,conventional protecting groups may be necessary to prevent certainfunctional groups from undergoing undesired reactions. The choice of asuitable protecting group for a particular functional group as well assuitable conditions for protection and deprotection are well known inthe art. For example, numerous protecting groups, and their introductionand removal, are described in T. W. Greene and G. M. Wuts, ProtectingGroups in Organic Synthesis, Second Edition, Wiley, New York, 1991, andreferences cited therein.

In a preferred method of synthesis, the α-aryl-N-alkylnitrone compoundsof this invention are prepared by coupling an aryl carbonyl compound offormula III:

wherein R¹-R⁴ are as defined above, with a hydroxylamine of formula IV:HO—NH—R⁵  IVwherein R⁵ is as defined above, under conventional reaction conditions.

The coupling reaction is typically conducted by contacting the arylcarbonyl compound III with at least one equivalent, preferably about 1.1to about 2 equivalents, of hydroxylamine IV in an inert polar solventsuch as methanol, ethanol, 1,4-dioxane, tetrahydrofuran, dimethylsulfoxide, dimethylformamide and the like. This reaction is preferablyconducted at a temperature of from about 0° C. to about 100° C. forabout 1 to about 48 hours. Optionally, a catalytic amount of an acid,such as hydrochloric acid, acetic acid, p-toluenesulfonic acid and thelike, may be employed in this reaction. Upon completion of the reaction,the α-aryl-N-alkylnitrone of formula I is recovered by conventionalmethods including precipitation, chromatography, filtration,distillation and the like.

The aryl carbonyl compounds of formula III employed in the couplingreaction are either known compounds or compounds that can be preparedfrom known compounds by conventional procedures. For example, suchcompounds are readily prepared by acylation of the corresponding arylcompound with the appropriate acyl halide under Friedel-Crafts acylationreaction conditions. Additionally, the formyl compounds, i.e. thosecompounds where 1 is hydrogen, can be prepared by formylation of thecorresponding aryl compound using, for example a disubstitutedformamides, such as N-methyl-N-phenylformamide, and phosphorousoxychloride (the Vilsmeier-Haack reaction), or using Zn(CN)₂ followed bywater (the Gatterman reaction). Numerous other methods are known in theart for preparing such aryl carbonyl compounds. Such methods aredescribed, for example, in I. T. Harrison and S. Harrison, Compendium ofOrganic Synthetic Methods, Wiley, N.Y., 1971, and references citedtherein.

Certain aryl carbonyl compounds of formula III can also be prepared byalkylation of the corresponding aryl hydroxy compound,(e.g.,4-hydroxybenzaldehyde and the like). This reaction is typicallyconducted by contacting the aryl hydroxy compound with a suitable base,such as an alkali or alkaline earth metal hydroxide, fluoride orcarbonate, in a inert solvent, such as ethanol, DMF and the like, todeprotonate the hydroxyl group. This reaction is generally conducted atabout 0° C. to about 50° C. for about 0.25 to 2 hours. The resultingintermediate is then reacted in situ with about 1.0 to about 2.0equivalents of an alkyl halide, preferably an alkyl bromide or iodide,at a temperature of from about 25° C. to about 100° C. for about 0.25 toabout 3 days.

Additionally, various aryl aldehydes of formula III can be prepared byreduction of the corresponding aryl nitrites. This reaction is typicallyconducted by contacting the aryl nitrile with about 1.0 to 1.5equivalents of a hydride reducing agent, such as LiAlH(OEt)₃, in aninert solvent such as diethyl ether, at a temperature ranging from about−78° to about 25° C. for about 1 to 6 hours. Standard work-up conditionsusing aqueous acid then provides the corresponding aryl aldehyde.

Preferred aryl carbonyl compounds include, but are not limited to,2-ethoxybenzaldehyde, 4-ethoxybenzaldehyde, 2-butoxybenzaldehyde,4-butoxybenzaldehyde, 4-pentyloxybenzaldehyde, 4-benzyloxybenzaldehyde,4-(4-fluorobenzyloxy)benzaldehyde, 4-(4-methoxybenzyloxy)benzaldehyde,4-hexyloxybenzaldehyde, 4-heptyloxybenzaldehyde,3-ethoxy-4-methoxybenzaldehyde, 4-ethoxy-3-methoxybenzaldehyde,3,4-diethoxybenzaldehyde, 3-ethoxy-4-hexyloxybenzaldehyde,2-fluoro-4-methoxybenzaldehyde, 2-fluoro-4-ethoxybenzaldehyde,2-fluoro-4-heptyloxybenzaldehyde, 2-fluoro-4-octyloxybenzaldehyde,4-benzyloxy-3-methoxybenzaldehyde, 4-phenoxy-3-methoxybenzaldehyde,3,4-methylenedioxybenzaldehyde (piperonal),3,4-ethylenedioxybenzaldehyde, 2,4,6-triethoxybenzaldehyde, and thelike.

The hydroxylamine compounds of formula V above are also known compoundsor compounds which can be prepared from known compounds by conventionalprocedures. Typically, the hydroxylamine compounds of formula V areprepared by reduction of the corresponding nitro compound (i.e., R⁵—NO₂,wherein R⁵ is as defined above) using a suitable reducing agent such asactivated zinc/acetic acid, activated zinc/ammonium chloride or analuminum/mercury amalgam. This reaction is typically conducted at atemperature ranging from about 15° C. to about 100° C. for about 0.5 to12 hours, preferably about 2 to 6 hours, in an aqueous reaction media,such as an alcohol/water mixture in the case of the zinc reagents or anether/water mixture in the case of the aluminum amalgams. Aliphaticnitro compounds (in the form of their salts) can also be reduced tohydroxylamines, using borane in tetrahydrofuran. Since somehydroxylamines have limited stability, such compounds are generallyprepared immediately prior to reaction with the aryl carbonyl compoundof formula III.

Preferred hydroxylamines for use in this invention include, but are notlimited to, N-cyclopentylhydroxyamine, N-tert-octylhydroxyamine,N-tert-butylhydroxylamine, N-isopropylhydroxylamine,N-n-propylhydroxylamine, N-n-butylhydroxylamine,N-tert-butylhydroxylamine, N-cyclohexylhydroxylamine,N-2,4-dimethyl-2-pentylhydroxylamine, 1-adamantylhydroxylamine and thelike.

Pharmaceutical Compositions

When employed as pharmaceuticals, the α-aryl-N-alkylnitrones of thisinvention are typically administered in the form of a pharmaceuticalcomposition. Such compositions can be prepared in a manner well known inthe pharmaceutical art and comprise at least one active compound.

Generally, the compounds of this invention are administered in apharmaceutically effective amount. The amount of the compound actuallyadministered will typically be determined by a physician, in the lightof the relevant circumstances, including the condition to be treated,the chosen route of administration, the actual compound administered,the age, weight, and response of the individual patient, the severity ofthe patient's symptoms, and the like.

The pharmaceutical compositions of this invention can be administered bya variety of routes including oral, rectal, transdermal, subcutaneous,intravenous, intra-muscular, and intranasal. Depending on the intendedroute of delivery, the compounds of this invention are preferablyformulated as either injectable or oral compositions.

The compositions for oral administration can take the form of bulkliquid solutions or suspensions, or bulk powders. More commonly,however, the compositions are presented in unit dosage forms tofacilitate accurate dosing. The term “unit dosage forms” refers tophysically discrete units suitable as unitary dosages for human subjectsand other, mammals, each unit containing a predetermined quantity ofactive material calculated to produce the desired therapeutic effect, inassociation with a suitable pharmaceutical excipient. Typical unitdosage forms include prefilled, premeasured ampules or syringes of theliquid compositions or pills, tablets, capsules or the like in the caseof solid compositions. In such compositions, the α-aryl-N-alkylnitronecompound is usually a minor component (from about 0.1 to about 50% byweight or preferably from about 1 to about 40% by weight) with theremainder being various vehicles or carriers and processing aids helpfulfor forming the desired dosing form.

Liquid forms suitable for oral administration may include a suitableaqueous or nonaqueous vehicle with buffers, suspending and dispensingagents, colorants, flavors and the like. Solid forms may include, forexample, any of the following ingredients, or compounds of a similarnature: a binder such as microcrystalline cellulose, gum tragacanth orgelatin; an excipient such as starch or lactose, a disintegrating agentsuch as alginic acid, Primogel, or corn starch; a lubricant such asmagnesium stearate; a glidant such as colloidal silicon dioxide; asweetening agent such as sucrose or saccharin; or a flavoring agent suchas peppermint, methyl salicylate, or orange flavoring.

Injectable compositions are typically based upon injectable sterilesaline or phosphate-buffered saline or other injectable carriers knownin the art. As before, the α-aryl-N-alkylnitrone compound in suchcompositions is typically a minor component, often being from about 0.05to 10% by weight with the remainder being the injectable carrier and thelike.

The above-described components for orally administrable or injectablecompositions are merely representative. Other materials as well asprocessing techniques and the like are set forth in Part 8 ofRemington's Pharmaceutical Sciences, 17th edition, 1985, Mack PublishingCompany, Easton, Pa., which is incorporated herein by reference.

The compounds of this invention can also be administered in sustainedrelease forms or from sustained release drug delivery systems. Adescription of representative sustained release materials can be foundin the incorporated materials in Remington's Pharmaceutical Sciences.

The following formulation examples illustrate representativepharmaceutical compositions of this invention. The present invention,however, is not limited to the following pharmaceutical compositions.

Formulation 1—Tablets

A compound of formula I is admixed as a dry powder with a dry gelatinbinder in an approximate 1:2 weight ratio. A minor amount of magnesiumstearate is added as a lubricant. The mixture is formed into 240-270 mgtablets (80-90 mg of active α-aryl-N-alkylnitrone compound per tablet)in a tablet press.

Formulation 2—Capsules

A compound of formula I is admixed as a dry powder with a starch diluentin an approximate 1:1 weight ratio. The mixture is filled into 250 mgcapsules (125 mg of active α-aryl-N-alkylnitrone compound per capsule).

Formulation 3—Liquid

A compound of formula I (125 mg), sucrose (1.75 g) and xanthan gum (4mg) are blended, passed through a No. 10 mesh U.S. sieve, and then mixedwith a previously made solution of microcrystalline cellulose and sodiumcarboxymethyl cellulose (11:89, 50 mg) in water. Sodium benzoate (10mg), flavor, and color are diluted with water and added with stirring.Sufficient water is then added to produce a total volume of 5 mL.

Formulation 4—Tablets

The compound of formula I is admixed as a dry powder with a dry gelatinbinder in an approximate 1:2 weight ratio. A minor amount of magnesiumstearate is added as a lubricant. The mixture is formed into 450-900 mgtablets (150-300 mg of active α-aryl-N-alkylnitrone compound) in atablet press.

Formulation 5—Injection

The compound of formula I is dissolved in a buffered sterile salineinjectable aqueous medium to a-concentration of approximately 5 mg/ml.

Utility

The α-aryl-N-alkylnitrones of this invention have been discovered toinhibit the formation of Aβ(142) beta-pleated sheets and/or protectagainst neuronal cell loss and/or inhibit the release of cytokines, suchas IL-1β and TNFα and/or protect against IL-1β/IFN_(γ)-induced toxicity.Additionally, such compounds have been found to reduce the cognitivedeficits caused by Aβ(25-35)/ibotenate as well as those developed bycertain autoimmune strains of mice. As previously discussed, theformation of Aβ(1-42) beta-pleated sheets, neuronal cell loss, betaamyloid-induced cognitive deficits are associated with neurodegenerativeconditions, such as Alzheimer's disease, and/or autoimmune conditions.Additionally, elevated levels of cytokines are associated withneurodegenerative, autoimmune and/or inflammatory conditions.Accordingly, the compounds and pharmaceutical compositions of thisinvention find use as therapeutics for preventing and/or treatingneurodegenerative, autoimmune and inflammatory conditions in mammalsincluding humans.

Surprisingly, it has also been discovered that the dimethoxy andtrimethoxy analogs of the compounds of formula I (i.e., compounds inwhich R¹ and R² are methoxy and R³ is hydrogen or R¹, R² and R³ are allmethoxy) have significantly higher toxicity than theα-aryl-N-alkylnitrone compounds of formula I. Due to their toxicity,such di- and trimethoxy compounds are not useful as therapeutic agentsor as analytical reagents for detecting free radicals in livingbiological systems.

Among the conditions which may be treated and/or prevented with theα-aryl-N-alkylnitrones of formula I are neurodegenerative conditions,such as Alzheimer's disease, Parkinson's disease, HIV-dementia and thelike; autoimmune conditions, such as systemic lupus, multiple sclerosisand the like; and inflammatory conditions, such as inflammatory boweldisease (IBD), rheumatoid arthritis, septic shock, erythema nodosumleprosy, septicemia, uveitis, adult respiratory distress syndrome (ARDS)and the like.

Additionally, since the α-aryl-N-alkylnitrones of this invention havebeen discovered to effectively inhibit the release of cytokines, such aIL-1β, IL-6 and TNFα, such compounds are useful for treating diseasescharacterized by an overproduction or a dysregulated production ofcytokines, particularly IL-1β, IL-6 and TNFα, including many autoimmuneand/or inflammatory conditions.

As discussed above, the compounds described herein are suitable for usein a variety of drug delivery systems. Injection dose levels fortreating neurodegenerative, autoimmune and inflammatory conditions rangefrom about 0.1 mg/kg/hour to at least 10 mg/kg/hour, all for from about1 to about 120 hours and especially 24 to 96 hours. A preloading bolusof from about 0.1 mg/kg to about 10 mg/kg or more may also beadministered to achieve adequate steady state levels. The maximum totaldose is not expected to exceed about 2 g/day for a 40 to 80 kg humanpatient.

For the prevention and/or treatment of long-term conditions, such asneurodegenerative and autoimmune conditions, the regimen for treatmentusually stretches over many months or years so oral dosing is preferredfor patient convenience and tolerance. With oral dosing, one to five andespecially two to four and typically three oral doses per day arerepresentative regimens. Using these dosing patterns, each dose providesfrom about 0.1 to about 20 mg/kg of the α-aryl-N-alkylnitrone, withpreferred doses each providing from about 0.1 to about 10 mg/kg andespecially about 1 to about 5. mg/kg.

When used to prevent the onset of a neurodegenerative, autoimmune orinflammatory condition, the α-aryl-N-alkylnitrones of this inventionwill be administered to a patient at risk for developing the condition,typically on the advice and under the supervision of a physician, at thedosage levels described above. Patients at risk for developing aparticular condition generally include those that have a family historyof the condition, or those who have been identified by genetic testingor screening to be particularly susceptible to developing the condition.

The compounds of this invention can be administered as the sole activeagent or they can be administered in combination with other agents,including other active α-aryl-N-alkylnitrone derivatives.

The novel α-aryl-N-alkylnitrones of this invention are also useful asanalytical reagents, i.e. as spin traps, for detecting unstable freeradicals using electron spin resonance (ESR) spectroscopy and relatedtechniques. When used as analytical reagents, the nitrone compounds ofthis invention are typically contacted with the radical to be studied insolution and an ESR spectrum generated in a conventional manner. Inparticular, the α-aryl-N-alkylnitrones of this invention may be used todetect and identify free radicals in biological systems. Any ESRspectrometer, such as a JEOL JES-FE3XG spectrometer, may be employed inthese experiments. Typically, the solution containing the spin-trap willbe deoxygenated by, for example, bubbling argon or nitrogen through thesolution before the ESR experiment is conducted. Preferably, an excessof the α-aryl-N-alkylnitrone is used in such ESR experiments.

The actual experimental procedures employed in the spin-trappingexperiment will depend on a number of factors, such as the manner ofradical production, the inertness of the solvent and reagents withrespect to the spin trap, the lifetime of the spin adduct and the like.Spin-trapping procedures are well known in the art and the exactprocedure employed can be determined by those skilled in the art.Typical procedures and apparatus for conducting spin trappingexperiments are described, for example, in C. A. Evans, “Spin Trapping”,Aldrichimica Acta, (1979), 12(2), 23-29, and references cited therein.

The following synthetic and biological examples are offered toillustrate this invention and are not to be construed in any way aslimiting the scope of this invention.

EXAMPLES

In the examples below, the following abbreviations have the followingmeanings. Abbreviations not defined below have their generally acceptedmeaning.

-   bd=broad doublet-   bs=broad singlet-   d=doublet-   dd=doublet of doublets-   dec=decomposed-   dH₂O=distilled water-   ELISA=enzyme-linked immuno-sorbent assay-   EtOAc=ethyl acetate-   EtOH=ethanol-   FBS=fetal bovine serum-   g=grams-   h=hours-   Hz=hertz-   IL-1β=interleukin-1β-   IL-6-=interleukin-6-   L=liter-   LPS=lipopolysaccharide-   m=multiplet-   min=minutes-   M=molar-   MeOH=methanol-   mg=milligram-   MHz=megahertz-   mL=milliliter-   mmol=millimole-   m.p.=melting point-   N=normal-   q=quartet-   quint.=quintet-   s=singlet-   t=triplet-   THF=tetrahydrofuran-   ThT=thioflavin T-   tic=thin layer chromatography-   TNFα=tumor necrosis factors-   μg=microgram-   μL=microliter-   UV=ultraviolet

In the examples below, all temperatures are in degrees Celsius (unlessotherwise indicated). Example A-C describe the synthesis ofintermediates useful for preparing α-aryl-N-alkylnitrones. The remainingexamples describe the synthesis of α-aryl-N-alkylnitrones of thisinvention and comparative α-aryl-N-alkylnitrones, and the ESR, in vitroand in vivo testing of such compounds.

Example A Synthesis of N-tert-Butylhydroxylamine

Zinc dust (648 g) was added in portions to a cooled mixture of2-methyl-2-nitropropane (503 g) and ammonium chloride (207 g) indeionized water (6 L) at such a rate so as to maintain the temperaturebelow 18° C. The reaction mixture was stirred mechanically for 15 hoursand then filtered. The solid was washed with hot water (1.75 L). Thecombined filtrate was saturated with potassium carbonate (4.6 Kg) andextracted with ethyl acetate (2×1300 mL). The organic solution was driedover anhydrous sodium sulfate, filtered and rotary evaporated to givethe title compound (329 g, 75.7% yield) as white crystals. This materialwas used without further purification.

Spectroscopic data were as follows:

¹H NMR (CDCl₃, 270 MHz) δ=1.090 (s, 3 CH₃).

Example B Synthesis of N-Isopropylhydroxylamine

Using the procedure of Example A above and 1-methyl-1-nitroethane, thetitle compound was prepared. The crude hydroxylamine product was usedwithout further purification.

Example C Synthesis of N-Cyclohexylhydroxylamine

Using the procedure of Example A above and 1-nitrocyclohexane, the titlecompound can be prepared. Alternatively, N-cyclohexylhydroxylaminehydrochloride may be purchased commercially from Aldrich ChemicalCompany, Inc., Milwaukee, Wis. USA and neutralized with a base, such aspotassium carbonate, to provide the title compound.

Example 1 Synthesis of α-(4-Heptyloxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 2 using 4-hydroxybenzaldehyde, 1-iodoheptane and2-methyl-2-nitropropane. The title compound was isolated in 60% overallyield as a solid, m.p. 68.5° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 3076.8 (CH), 2972.3 (CH), 1601.9 (C═N), 1250.9 (C—O—C)and 1118.8 (N—O).

¹H NMR (CDCl₃, 270 MHz)=8.25 (2H, d, J=8.9 Hz, phenyl 2H), 7.44 (1H, s,nitronyl H), 6.90 (2H, d, J=8.9 Hz, phenyl 2H), 3.98 (2H, t, J=6.7 Hz,CH₂), 1.77 (2H, quintet, J=6.7 Hz, CH₂), 1.58 (9H, s, 3 CH₃), 1.36 (8H,m, 4 CH₂) and 0.87 (3H, t, J=6.7 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz) δ=160.9, 131.0, 129.8, 124.0, 114.4, 69.9,68.0, 31.5, 28.9, 28.7, 28.0, 25.6, 22.3 and 13.7.

Example 2 Synthesis of α-(4-Hexyloxyphenyl)-N-n-propylnitrone

A solution of 4-hydroxybenzaldehyde (27.11 g, 0.222 moles) in ethanolwas refluxed with sodium hydroxide (8.88 g, 0.222 moles) for 30 minutes.1-Iodohexane (47.10 g, 0.222 moles) was added in one portion and thesolution refluxed for 68 hours. The ethanol was removed by rotaryevaporation and the residue was reacted with 1-nitropropane, ammoniumchloride, and zinc dust in H₂O/ethanol (300:20, v:v) for 18 hours atroom temperature. The reaction mixture was filtered, the solvent removedby rotary evaporation, and the residue purified by column chromatographyusing ethyl acetate/hexane (1:1, v:v) as the eluant (R_(f)=0.42 on asilica gel plate using ethyl acetate/hexane (1:1, v:v) as the eluant).The title compound was isolated as a solid (1.63 g, 12.4% overallyield), m.p. 45° C.

Spectroscopic data was as follows:

¹H NMR (CDCl₃, 270 MHz): δ=8.22 (2H, d, J=8.8 Hz, phenyl 2H), 7.28 (1H,s, nitronyl H), 6.92 (2H, d, J=8.8 Hz, phenyl 2H), 3.93 (4H, m, 2CH₂),2.07 (4H, m, CH₂), 1.36 (6H, m, 3CH₂), 1.00 (3H, t, CH₃), 0.908 (3H, t,CH₃).

Examples 3-6

Using the procedures described herein, the following compounds wereprepared:

-   -   α-(3-Ethoxy-4-methoxyphenyl)-N-tert-butylnitrone    -   α-(4-Ethoxyphenyl)-N-tert-butylnitrone    -   α-(4-Benzyloxy-3-methoxyphenyl)-N-tert-butylnitrone, and    -   α-[3-(4-Methoxyphenoxy)phenyl]-N-tert-butylnitrone.

Example 7 Synthesis of α-(2-Ethoxyphenyl)-N-tert-butylnitrone

2-Ethoxybenzaldehyde (12.0 g, 79.90 mmol) and N-tert-butylhydroxylamine(10.69 g, 119.86 mmol) were mixed in chloroform with molecular sieves(50 g, 4 A) and silica gel (10 g). The mixture was sealed under argongas and stirred for 70 h at room temperature. The mixture was thenfiltered and the solid washed with ethyl acetate and the combinedsolution was rotary evaporated. Pentane (50 mL) was added to the liquidresidue and isolation of the resulting solid afforded 13.79 g (78.0%yield) of the title compound as white crystals, m.p. 58.3° C.(R_(f)=0.55 on a silica gel plate using ethyl acetate as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2976.7 (CH), 2935 (CH), 1597.0 (C═N), 1567.1 (benzenering) and 1123.6 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=9.322 (1H, dd, J₁=1.7 Hz, J₂=7.9 Hz, phenylH), 8.067 (1H, s, CH═N), 7.302 (1H, td, J_(t)=7.9 Hz, J_(d)=1.7 Hz,phenyl H), 6.979 (1H, td, J_(t)=7.9 Hz, J_(d)=0.5 Hz, phenyl H), 6.839(1H, d, J=7.9 Hz, phenyl H), 4.055 (2H, q, J=6.9 Hz, OCH₂), 1.586 (9H,s, 3 CH₃) and 1.423 (3H, t, J=6.9 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=156.948, 131.323, 128.836, 124.688,120.813, 120.386, 110.868, 70.767, 63.842, 27.997 and 14.375.

Examples 8-10

Using the procedures described herein, the following compounds wereprepared:

-   -   α-(3,4-Ethylenedioxyphenyl)-N-tert-butylnitrone, and    -   α-(3,4-Methylenedioxyphenyl)-N-tert-butylnitrone.    -   α-(4-ethoxyphenyl)-N-n-butylnitrone

Example 11 Synthesis of α-(Ethoxyphenyl)-N-cyclohexylnitrone

A solution of 4-ethoxybenzaldehyde (6.62 g, 44.1 mmol) in 200 mL ofbenzene was refluxed with N-cyclohexylhydroxylamine (6.61 g, 57.4 mmol)in the presence of p-toluenesulfonic acid (0.8 g, 4 mmol) for 72 h.After rotary evaporation, the residue was purified by, recrystallizationfrom hexanes and ethylene glycol dimethyl ether (100 mL, 3:1, v:v) togive the title compound (9.2 g, 84% yield) as a solid, m.p. 124.0° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹); 2933.0, 2862 (CH), 1599.6 (C═N), 1297.0 (C—O—C) and1149.4 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.20 (2H, d, J=8.9 Hz, phenyl 2H), 7.32 (1H,s, nitronyl H), 6.88 (2H, d, J=8.9 Hz, phenyl 2H), 4.05 (2H, quartet,J=7.0 Hz, CH₂), 3.75 (1H, m, CH), 1.94 (6H, m, 6 CH), 1.68 (2H, m, 2CH), 1.39 (2H, t, J=7.0 Hz, CH₃) and 1.27 (2H, m, 2 CH).

¹³C NMR (CDCl₃, 67.9 MHz): δ=160.6, 132.1, 130.7, 123.8, 114.4, 75.0,63.4, 30.8, 24.7 and 14.3.

Example 12 Synthesis ofα-(4-Benzyloxy-3-methoxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 11 using 4-benzyloxy-3-methoxybenzaldehyde andN-cyclohexylhydroxylamine. The title compound was isolated in 97.9%yield as a solid, m.p. 154.1° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2935.3 (CH), 1595.4 (C═N), 1265.1 (C—O—C) and 1147.6(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.50 (1H, d, J=2.0 Hz, phenyl H), 7.34 (7H,m, phenyl H & nitronyl H), 6.86 (1H, d, J=8.4 Hz, phenyl H), 5.19 (2H,s, CH₂), 3.94 (3H, s, CH₃), 3.78 (1H, m, cyclohexyl H), 1.95 (6H, m, 6cyclohexyl H), 1.67 (2H, m, 2 CH) and 1.30 (2H, m, 2 CH).

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.0, 149.4, 137.0, 132.5, 128.8, 128.2,127.4, 124.8, 122.9, 113.3, 111.6, 75.2, 70.7, 55.7, 30.8 and 24.7.

Example 13 Synthesis of α-(3-Ethoxy-4methoxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the, procedure described inExample 11 using 3-ethoxy-4-methoxybenzaldehyde andN-cyclohexylhydroxylamine. The title compound was isolated in 57% yieldas a solid, m.p. 113.5° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹) 2857.3. (CH), 1590.8 (C═N), 1265.0, 1239.0 (C—O—C) and1126.1 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.42 (1H, d, J=1.8 Hz, phenyl H), 7.39 (1H,dd, J=7.5 and 1.8 Hz, phenyl H), 7.32 (1H, s, nitronyl H), 6.84 (1H, d,J=7.5 Hz, phenyl H), 4.14 (2H, quartet, J=7.0 Hz, CH₂), 3.88 (3H, s,CH₃), 3.76 (1H, m, CH), 1.96 (6H, m, 6 CH), 1.68 (1H, m, CH), 1.44 (3H,t, J=7.0 Hz, CH₃) and 1.27 (3H, m, 3 CH).

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.95, 148.13, 132.47, 124.32, 122.93,112.36, 110.91, 75.14, 64.10, 55.71, 30.80, 24.75 and 14.34.

Example 14 Synthesis of α-(3,4Ethylenedioxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 11 using 3,4-ethylenedioxybenzaldehyde andN-cyclohexylhydroxylamine. The title compound was isolated in 74.5,%yield as a solid, m.p. 96.7° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2927.9 (CH), 1575.6 (C═N), 1319.5 (C—O—C) and 1133.9(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=7.98 (1H, d, J=2.0 Hz, phenyl H), 7.60 (1H,dd, J=7.4 & 2.0 Hz, phenyl H), 7.27 (1H, s, nitronyl H), 6.83 (1H, d,J=7.4 Hz, phenyl H), 4.24 (4H, m, 2 CH₂), 3.75 (1H, m, CH), 1.94 (7H, m,7 CH) and 1.28 (3H, m, 3 CH).

¹³C NMR (CDCl₃, 67.9 MHz): δ=145.45, 143.45, 131.86, 124.75, 122.98,117.76, 117.26, 75.16, 64.50, 63.98, 30.80 and 24.73.

Example 15

Using the procedures described herein, the following compound wasprepared:

-   -   α-(4-Ethoxy-3-methoxyphenyl)-N-cyclohexylnitrone.

Example 16 Synthesis of α-(3,4-Ethylenedioxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedure described inExample 11 using 3,4-ethylenedioxybenzaldehyde andN-isopropylhydroxylamine. The crude produce was purified by columnchromatography over silica gel using ethyl acetate as the eluant. Thetitle compound was isolated in 53% yield as a solid, m.p. 108.8° C.(R_(f)=0.31 on a silica gel plate using EtOAc as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2978.9 (CH), 1582.3 (C═N), 1297.0 (C—O—C) and 1063.8(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=7.99 (1H, d, J=2.0 Hz, phenyl H), 7.61 (1H,dd, J=8.5 & 2.0 Hz, phenyl H), 7.28 (1H, s, nitronyl H), 6.84 (1H, d,J=8.5 Hz, phenyl H), 4.25 (4H, m, 2 CH₂), 4.13 (1H, septet, J=6.7 Hz,CH) and 1.46 (6H, d, J=6.7 Hz, 2 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=145.5, 143.5, 131.6, 124.7, 123.0, 117.8,117.3, 67.3, 64.5, 64.0 and 20.5.

Example 17 Synthesis of α-(3-Ethoxy-4-methoxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedures described inExamples 11 using 3-ethoxy-4-methoxybenzaldehyde andN-isopropylhydroxylamine. The title compound was isolated in 43.9% yieldas a solid, m.p. 80.8° C. (R_(f)=0.15 on a silica gel plate using ethylacetate as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2981.6 (CH), 1596.7 (C═N), 1443.7 (CH₃), 1263.3 (C—O—C)and 1128.6 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.44 (1H, d, J=1.9 Hz, phenyl H), 7.40 (1H,dd, J=8.5 & 1.9 Hz, phenyl H), 7.34 (1H, s, nitronyl CH), 6.87 (1H, d,J=8.5 Hz, phenyl H), 4.16 (3H, m, Cl and CH), 3.89 (3H, s, CH₃) and 1.48(9H, m, 3 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=151.0, 148.2, 132.2, 124.2, 123.0, 112.3,110.9, 67.2, 64.1, 55.7, 20 5 and 14.4.

Example 18 Synthesis of α-(2-Ethoxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedure described inExample 11 using 2-ethoxybenzaldehyde and N-isopropylhydroxylamine. Thetide compound was isolated in 48.8% yield as a solid, m.p. 59.4° C.(R_(f)=0.48 on a silica gel plate using ethyl acetate as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2978.8 (CH), 1593.6 (C═N), 1245.0 (C—O—C) and 1149.3(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=9.30 (1H, d, J=7.7 Hz, phenyl H), 7.961 (1H,s, nitronyl H), 7.30 (1H, td, J=7.7 & 1.7 Hz, phenyl H), 6.98 (1H, td,J=7.7 &1.7 Hz, phenyl H), 6.83 (1H, d, J=7.7 Hz, phenyl H), 4.23 (1H, m,CH), 4.03 (2H, quartet, J=7.2 Hz, CH₂) and 1.44 (9H, m, 3 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=156.56, 131.38, 129.02, 126.76, 120.80,120.04, 110.75, 67.95, 63.78, 20.55 and 14.39.

Example 19 Synthesis of α-(2-Ethoxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 11 using 2-ethoxybenzaldehyde and N-cyclohexylhydroxylamine. Thetitle compound was isolated in 89% yield as a solid, m.p. 54.8° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2932.9 (CH), 1593.8 (C═N), 1244.9 (C—O—C) and 1144.8(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=9.32 (1H, d, J=7.9 Hz, phenyl H), 7.89 (1H,s, nitronyl H), 7.29 (1H, t, J=7.9 Hz, phenyl H), 6.97 (1H, t, J=7.9 Hz,phenyl H), 6.84 (1H, d, J=7.9 Hz, phenyl H), 4.06 (2H, quartet, J=7.1Hz, CH₂), 3.84 (1H, m, CH), 1.95 (6H, m, 2 CH₂ & 2 CH), 1.67 (1H, m,CH), 1.66 (3H, t, J=7.1.Hz, CH₃) and 1.25 (3H, m, 3 CH).

¹³C NMR (CDCl₃, 67.9 MHz): δ=156.6, 131.3, 129.0, 127.1, 120.8, 120.1,110.7, 75.8, 63.8, 30.8, 24.7 and 14.4.

Example 20 Synthesis ofα-(4-Benzyloxy-3-methoxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedures described inExamples 11 using 4-benzyloxy-3-methoxybenzaldehyde andN-isopropylhydroxylamine. The title compound was isolated in 54.6% yieldas a solid, m.p. 95.5° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2988.4 (CH), 2935.0 (CH), 1585.1 (C═N), 1461.0 (CH₃),1262.9 (C—O—C) and 1126.9 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.50 (1H, d, J=1.7 Hz, phenyl H), 7.33 (7H,m, 6 phenyl H & nitronyl H), 6.86 (1H, d, J=8.4 Hz, phenyl H), 5.18 (2H,s, CH₂), 4.13 (1H, septet, J=6.4 Hz, CH), 3.93 (3H, s, CH₃) and 1.47(6H, d, J=6.4 Hz, 2 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=149.8, 149.4, 136.9, 132.2, 128.8, 128.2,127.4, 124.6, 122.9, 113.2, 111.5, 70.7, 67.3, 55.7 and 20.5.

Example 21

Using the procedures described herein, the following compound wasprepared:

-   -   α-(4-Ethoxy-3-methoxyphenyl)-N-isopropylnitrone.

Example 22 Synthesis ofα-(3-Ethoxy-4-hexyloxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 28 using 3-ethoxy-4-hydroxybenzaldehyde, 1-iodohexane andN-cyclohexylhydroxylamine. The title compound was isolated in 41.3%yield as a solid, m.p. 67.3° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2920.7 (CH), 1597.7 (C═N), 1341.2 (CH₃), 1267.7 (C—O—C),and 1129.0 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.36 (1H, d, J=1.9 Hz, phenyl H), 7.39 (1H,dd, J=8.6 & 1.9 Hz, phenyl H), 7.31 (1H, s, nitronyl H), 6.84 (1H, d,J=8.6 Hz, phenyl H), 4.12 (2H, quartet, J=7.0 Hz, CH₂), 4.01 (2H, t,J=6.8 Hz, CH₂), 3.76 (1H, m, CH), 1.93 (10H, m, 5 CH₂), 1.42 (3H, t,J=7.0 Hz, CH₃), 1.32 (8H, m, 4 CH₂) and 0.88 (3H, t, J=7.0 Hz, CH₃).

13C NMR (CDCl₃, 67.9 MHz): δ=150.8, 157.6, 132.6, 124.2, 123.0, 113.2,112.6, 75.1, 69.0, 64.4, 31.3, 30.8, 28.7, 25.3, 24.8, 22.2, 14.4 and13.6.

Example 23 Synthesis of α-(4Benzyloxy-3-methoxyphenyl)-N-n-butylnitrone

The title compound was prepared according to the procedures described inExamples 11 using 4-benzyloxy-3-methoxybenzaldehyde andN-n-butylhydroxylamine. The title-compound was isolated in 41.7% yieldas a solid, m.p. 81.2° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2925.1 (CH), 2856.9 (CH), 1593.2 (C═N), 1463.1 (CH₃),1263.1 (C—O—C) and 1156.1 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.42 (1H, d, J=2.0 Hz, phenyl H), 7.34 (7H,m, 6 phenyl H & nitronyl H), 6.86 (1H, d, J=8.4 Hz, phenyl H), 5.18 (2H,s, CH₂), 3.93 (3H, s, CH₃), 3.93 (2H, t, J=7.3 Hz, CH₂), 1.96 (2H,quintet, J=7.3 Hz, CH₂), 1.39 (2H, sextet, J=7.3 Hz, CH₂) and 0.95 (3H,t, J=7.3 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.0, 149.4, 136.9, 134.3, 128.8, 128.2,127.4, 124.4, 122.9, 113.2, 111.4, 70.7, 66.6, 55.8, 29.4, 19.4 and13.2.

Example 24

Using the procedures described herein, the following compound wasprepared:

-   -   α-(4-Ethoxy-3-methoxyphenyl)-N-n-butylnitrone.

Example 25 Synthesis of α-(2-Ethoxyphenyl)-N-n-butylnitrone

The title compound was prepared according to the procedures described inExamples 11 using 2-ethoxybenzaldehyde and N-n-butylhydroxylamine. Thetitle compound was isolated in 44.5 % yield as a liquid.

Spectroscopic data were as follows:

IR (NaCl, cm⁻¹): 2959.6 (CH), 1594.9 (C═N), 1454.8 (CH₃), 1245.1 (C—O—C)and 11.63.5 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=9.24 (1H, d, J=8.0 Hz, phenyl H), 7.80 (1H,s, nitronyl H), 7.28 (1H, t, J=8.0 Hz, phenyl H), 6.95 (1H, t, J=8.0 Hz,phenyl H), 6.81 (1H, d, J=8.0 Hz, phenyl H), 4.02 (2H, quartet, J=6.35Hz, CH₂), 3.90 (2H, t, J=7.1 Hz, CH₂), 1.93 (2H, quintet, J=7.3 Hz,CH₂), 1.40 (5H, m, CH₂ & CH₃) and 0.93 (3H, t, J=7.4 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=156.6, 131.5, 129.0, 128.9, 120.7, 119.9,110.8, 67.2, 63.8, 29.5, 19.3, 14.3 and 13.2.

Example 26 Synthesis of α-(3-Ethoxy-4-methoxyphenyl)-N-n-butylnitrone

The title compound was prepared according to the procedures described inExamples 11 using 3-ethoxy-4-methoxybenzaldehyde andN-n-butylhydroxylamine. The title compound was isolated in 41.1% yieldas a solid, m.p. 117.3° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2953.1 (CH), 1593.9 (C═N), 1265.4 (C—O—C) and 1129.3(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.35 (1H, d, J=1.8 Hz, phenyl H), 7.42 (1H,dd, J=8.5 & 1.8 Hz, phenyl H), 7.27 (1H, s, nitronyl H), 6.86 (1H, d,J=8.5 Hz, phenyl H), 4.16 (2H, quartet, J=6.9 Hz, CR₂), 3.87 (5H, m, CH₂and CH₃), 1.94 (2H, quintet, J=7.4 Hz, CH₂), 1.45 (5H, m, CH₂ and CH₃)and 0.95 (3H, t, J=7.4 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=151.2, 148.2, 132.3, 124.1, 122.9, 112.3,111.0, 66.6, 64.2, 55.7, 29.4, 19.4, 14.3 and 13.2.

Example 27 Synthesis of α-(3-Ethoxy-4-hexyloxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedure described inExample 28 using 3ethoxy-4-hydroxybenzaldehyde, 1-iodohexane andN-isopropylhydroxylamine. The title compound was isolated in 47.1%overall yield as a solid, m.p. 69.0° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2995.0 (CH), 1596.9 (C═N), 1393.8 (iPr), 1261.2 (C—O—C)and 1128.7 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.36 (1H, d, J=2.0 Hz, phenyl H), 7.40 (1H,dd, J=8.4 & 2.0 Hz, phenyl H), 7.32 (1H, s, nitronyl H), 6.86 (1H, d,J=8.4 Hz, phenyl H), 4.13 (3H, m, CH₂ and CH), 4.02 (2H, t, J=6.9 Hz,CH₂), 1.82 (2H, quintet, J=7.4 Hz, CH₂), 1.48 (6H, d, J=6.7 Hz, 2 CH₃),1.42 (3H, t, J=6.9 Hz, CH₃), 1.31 (6H, m, 3 CH) and 0.88 (3H, t, J=6.9Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.8, 148.6, 132.2, 124.1, 123.1, 133.2,112.6, 69.0, 67.2, 64.4, 31.3, 28.7, 25.3, 22.2, 20.5, 14.4 and 13.6.

Example 28 Synthesis ofα-(3-Ethoxy-4-hexyloxyphenyl)-N-tert-butylnitrone

A solution of 3-ethoxy-4-hydroxybenzaldehyde (13.28 g, 79.9 mmol) andsodium hydroxide. (3.20 g, 79.9 mmol) in ethanol (120 mL) was refluxedfor 30 min. To the refluxing solution was added 1-iodohexane (18.6 g,87.9 mmol) in one portion and reflux was continued for 24 h. Thesolution was then cooled and the ethanol removed on a rotary evaporator.The residue was dissolved in ethyl acetate and this solution filteredand rotary evaporated. The resulting residue was reacted withN-tert-butylhydroxylamine (6.94 g) in 200 mL of benzene in the presenceof p-toluenesulfonic acid (0.8 g) at refluxing temperature for 24 h.After evaporation, the residue obtained was purified byrecrystallization from hexanes to give the title compound (11.02 g,57.2% overall yield) as a solid, m.p. 35.5° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2900 (CH), 1596.2 (C═N), 1361.1 (CH₃), 1276.0 (C—O—C)and 1144.8 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.38 (1H, d, J=1.7 Hz, phenyl H), 7.45 (1H,dd, J=8.5 & 1.7 Hz, phenyl H), 7.42 (1H, s, nitronyl H), 6.86 (1H, d,J=8.5 Hz, phenyl H), 4.13 (2H, quartet, J=7.0 Hz, CH₂), 4.02 (2H, t,J=6.8 Hz, CH₂), 1.82 (2H, m, CH₂), 1.65 (2H, m, CH₂), 1.58 (9H, s, 3CH₃), 1.42 (3H, t, J=7.0 Hz, CH₃), 1.31 (4H, m, 2 CH₂) and 0.88 (3H, t,J=6.3 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.8, 148.6, 130.1, 124.4, 123.4, 113.4,112.6, 70.0, 69.0, 64.4, 31.3, 28.7, 28.0, 25.3, 22.2, 14.4 and 13.6.

Example 29 Synthesis ofα-(2-Fluoro-4-octyloxyphenyl)-N-tert-butylnitrone

To dry dimethylformamide (200 mL) were added2-fluoro-4-hydroxybenzonitrile (13.71 g, 100 mmol), 1-iodooctane (28.82g, 120 mmol) and potassium fluoride (11.6 g, 200 mmol). This mixture wasstirred at room temperature for 16 h, and then at 50° C. for 2 h andthen at 90° C. for 2 h. The mixture was then poured into wet-ice (400 g)and 37% HCl (10 mL). The resulting solution was extracted with diethylether (3×200 mL). The ether layer was washed with water (2×200 mL) anddried over Na₂SO₄. After filtration, rotary evaporation gave the crudedesired intermediate 2-fluoro-4-n-octyloxybenzonitrile (27.83 g). Thisliquid intermediate was then added, over a 5-10 min period at 3-13° C.,to a flask containing LiAlH(OEt)₃ [which had been freshly prepared fromLiAH₄ (5.03 g, 0.1326 mol) and ethyl acetate (15.24 g, 0.1730 mol) at3-8° C. in diethyl ether (130 mL)]. The reaction mixture was stirred at5° C. for 75 min and 5 N H₂SO₄ aqueous solution (120 mL) was addeddropwise with cooling. After separation, the aqueous layer was extractedwith diethyl ether (2×100 mL) and the combination extracts were Washedwith water (2×100 mL). Standard work-up procedures afforded crude2-fluora-4-n-octyloxybenzaldehyde (26.07 g). The crude material was thenmixed with N-tert-butylhydroxylamine (8.6 g, 96.4 mmol), molecularsieves (50 g, 4 A) and silica gel (10 g) in chloroform (250 mL). Themixture was stirred at, room temperature for 23 h and refluxed for 3 hunder argon gas. The mixture was then filtered and rotary evaporated togive a residue which was purified by column chromatography over silicagel eluted with he hexanes/ethyl acetate (4:1, v:v). The title compound(12.90 g) was obtained in 39.9% overall yield as a slightly yellowishsolid, m.p. 35.6° C. (R_(f)=0.36 on a silica gel plate usinghexanes/EtOAc, 4:1, v:v, as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2928.1 (CH), 2855:1 (CH), 1617.8 (C═N), 1556.8 (benzenering), 1287.0 (Ar—F), 1161.2 (Ar—O), 1129.4 (N—O) and 1105.4 (alklyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=9.288 (1H, t, J_(H)=J_(F)=8.9 Hz, aromaticH), 7.702 (1H, s, CH═N), 6.684 (1H, dd, J_(H)=8.9 Hz, J_(H)=2.5 Hz,aromatic H), 6.586 (1 H, dd, J_(F)=13.7 Hz, J_(H)=2.5 Hz, aromatic H),3.937 (2H, t, J=6.6 Hz, OCH₂), 1.745 (2H, m, CH₂),1.568 (9H, s, 3 CH₃),1.408-1.251 (10H, m, (CH₂)₅) and 0.851 (3H, t, J=6.9 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=161.959 (d, J=11.4 Hz), 161.959 (d, J=253.9Hz), 130.103, 122.232 (d, J=8.3 Hz), 112.363 (d, J=8.3 Hz), 109.892;101.693 (d, J=25.9 Hz), 70.630, 68.372, 31.459, 28.958, 28.851, 28.683,27.936, 25.587, 22.261 and 13.658.

Example 30 Synthesis of α-(2,4,6-Triethoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 11 using 2,4,6-triethoxybenzaldehyde andN-tert-butylhydroxylamine. The title compound was isolated in 92.3%yield as a solid, m.p. 109.1° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2978.5 (CH), 1608.2 (C═N), 1438.6 (CH₃), 1231.2 (C—O—C)and 1132.3 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=7.46 (1H, s, nitronyl H), 6.07 (2H, s, 2phenyl H), 3.98 (6H, m, 3 CH₂), 1.56 (9H, s, 3 CH₃) and 1.32 (9H, m, 3CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=161.9, 159.3, 125.0, 92.3, 69.3, 63.9,63.4, 28.1, 14.5 and 14.3.

Example 31 Synthesis of α-(2,4,6-Triethoxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 11 using 2,4,6-triethoxybenzaldehyde andN-cyclohexylhydroxylamine.

The title compound was isolated in 87.4% yield as a solid, m.p. 145.7°C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2935 (CH), 1601 (C═N), 1391 (CH₃), 1167 (C—O—C) and 1133(N—O).

¹H NMR (CDCl₃, 270 MHz): δ=7.34 (1H, s, nitronyl H), 6.06 (2H, s, 2phenyl H), 3.99 (6H, m, 3 CH₂), 3.80 (1H, m, CH), 1.94 (1OH, m, 5 CH₂)and 1.32 (9H, m, 3 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=162.1, 159.4, 127.2, 102.0, 92.2, 74.1,64.0, 63.4, 31.0, 29.6, 24.8, 14.5 and 14.4.

Example 32 Synthesis of α-(2-n-Butoxyphenyl)-N-tert-butylnitrone

The title compound-was prepared according to the procedure described inExample 28 using 2-hydroxybenzonitrile, 1-iodobutane andN-tert-butylhydroxylamine. The title compound was isolated in 77.4%overall yield as a viscous oil.

Spectroscopic data were as follows:

IR (NaCl, cm⁻¹): 3074 (Ar CH), 2962 (CH), 1594 (C═N), 1468 (CH₃), 1244(C—O—C) and 1132 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=9.29 (1H, dd, J=7.9 & 1.7 Hz, phenyl H), 8.06(1H, s, nitronyl H), 7.29 (1H, td, J=7.9 & 1.7 Hz, phenyl H), 6.96 (1H,t, J=7.9 Hz, phenyl H), 6.82 (1H, d, J=7.9 Hz, phenyl H), 3.98 (2H, t,J=6.3 Hz, CH₂), 1.75 (2H, quintet, J=6.9 Hz, CH₂), 1.57 (9H, m, 3 CH₃),1.50 (2H, m, CH₂) and 0.96 (3H, t, J=7.3 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=157.1, 131.4, 128.8, 124.7, 120.7, 120.4,110.8, 70.7, 67.9, 30.9, 28.0, 19.0 and 13.4.

Example 33 Synthesis of α-(3,4-Diethoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 11 using 3,4-diethoxybenzaldehyde and N-tert-butylhydroxylamine.The title compound was isolated in 93.7% yield as a solid, m.p. 57.9° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2984 (CH), 1596 (C═N), 1272 (C—O—C) and 1146 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.41 (1H, d, J=1.9 Hz, phenyl H), 7.46 (1H,dd, J=8.4 & 1.9 Hz, phenyl H), 7.43 (1H, s, nitronyl H), 6.86 (1H, d,J=8.4 Hz, phenyl H), 4.14 (2H, quartet, J=7.0 Hz, CH₂), 4.13 (2H,quartet, J=7.0 Hz, CH₂), 1.58 (9H, s, 3 CH₃), 1.45 (3H, t, J=7.0 Hz,CH₃) and 1.44 (3H, t, J=7.0 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz):=150.5, 148.4, 130.2, 124.4, 123.4, 113.0,112.3, 70.1, 64.3, 28.0, 14.4 and 14.3.

Example 34 Synthesis ofα-(2-Fluoro-4-heptyloxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 28 using 2-fluoro-4-hydroxybenzonitrile, 1-iodoheptane andN-tert-butylhydroxylamine. The title compound was isolated in 66.0%overall yield as a white solid, m.p. 38.8° C. (R_(f)=0.21 on a silicagel plate using hexanes/ethyl acetate, 4:1, v:v, as the eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2930.1 (CH), 2857.5 (CH), 1618.6 (C═N), 1556.6 (enzenering), 1286.8 (Ar—F), 1161.6 (Ar—O), 1129.4 (N—O) and 1105.4 (alkyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=9.291 (1H, t, J_(H)=J_(F)=8.9 Hz, aromaticH), 7.723 (1H, s, CH═N), 6.700 (1H, dd, J_(H)=8.9 Hz, J_(H)=2.6 Hz,aromatic H), 6.603 (1H, dd, J_(F)=13.1 Hz, J_(H)=2.6 Hz, aromatic H),3.954 (2H, t, J=6.6 Hz, OCH₂), 1.747 (2H, m, CH₂), 1.585 (9H, s, 3 CH₃),1.445-1.286 (8 H, m, (CH₂)₄) and 0.872 (3H, t, J=6.8 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=161.997 (d, J=12.4 Hz), 161.990 (d, J=253.9Hz), 130.133, 122.354 (d, J=8.3 Hz), 112.332 (d, J=8.3 Hz), 109.922,101.716 (d, J=24.9 Hz), 70.645, 68.387, 31.429, 28.683, 27.951, 25.556,22.216 and 13.658.

Example 35 Synthesis of α-(2-Fluoro-4-ethoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 29 using 2-fluoro-4-hydroxybenzonitrile, ethyl iodide andN-tert-butylhydroxylamine. The title compound was isolated in 64.7%overall yield as slightly yellowish crystals, m.p. 82.5° C. (R_(f)=0.16on a silica gel plate using hexanes/ethyl acetate, 4:1, v:v, as theeluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2978.4 (CH), 2938.0 (CH), 1616.3 (C═N), 1560.7 (benzenering), 1290.0 (Ar—O), 1128.7 (N—O), 1112.9 (Ar—F) and 1042.3 (alkyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=9.300 (1H, t, J=9.0 Hz, aromatic H), 7.716(1H, s, nitronyl CH), 6.695 (1H, dd, J=9.0 Hz, J=2.4 Hz, aromatic H),6.597 (1H, dd, J=13.1 Hz, J=2.4 Hz, aromatic H), 4.035 (2H, q, J=6.9 Hz,OCH₂), 1.581 (9H, s, 3 CH₃) and 1.396 (3H, t, J=6.9 Hz, CH₃).

¹³C-NMR (CDCl₃, 67.9 MHz): δ=161.952 (d, J_(F)=254.9 Hz); 161.738 (d,J_(F)=12.4 Hz), 130.118, 122.285 (d, J_(F)=9.3 Hz), 112.409 (d,J_(F)=8.3 Hz), 109.846, 101.708 (d, J_(F)=25.9 Hz), 70.660, 63.842,27.936 and 14.177.

Example 36 Synthesis of α-(2-Fluoro-4-ethoxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 29 using 2-fluoro-4-hydroxybenzonitrile, ethyl iodide andN-cyclohexylhydroxylamine. The title compound was isolated in 58.8%overall yield as slightly yellowish crystals, m.p. 112.7° C. (R_(f)=0.17on a silica gel plate using hexanes/ethyl acetate, 4:1, v:v, as theeluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2956.5 (CH), 2933.2 (CH), 1616.9 (C═N), 1558.7 (benzenering), 1287.4 (Ar—O), 1158.7 (N—O), 1103.5 (Ar—F) and 1039.6 (alkyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=9.245 (1H, t, J=9.0 Hz, aromatic H), 7.580(1H, s, nitronyl CH), 6.689 (1H, dd, J=9.0 Hz, J=2.5 Hz, aromatic H),6.580 (1H, dd, J=14.3 Hz, J=2.5 Hz, aromatic H), 4.022 (2H, q, J=6.9 Hz,OCH₂), 3.805 (1H, tt, J=11.3 Hz, J=4.1 Hz, N—CH), 2.069-1.990 (2H, m,cyclohexyl 2H), 1.958-1.862 (4H, m, cyclohexyl 4H), 1.694-1.651 (1H, m,cyclohexyl H), 1.386 (3H, t, J=6.9 Hz, CH₃) and 1.333-1.176 (3H, m,cyclohexyl 3H).

¹³C NMR (CDCl₃, 67.9 MHz): δ=161.734 (d, J_(F)=13.0 Hz), 161.639 (d,J_(F)=253.9 Hz), 130.255, 124.703 (d. J_(F)=8.3 Hz), 112.165 (d,J_(F)=8.3 Hz), 109.953, 101.731 (d, J_(F)=24.9 Hz), 75.587, 63.857,30.819, 24.702 and 14.177.

Examples 37-38

Using the procedures described herein, the following compounds wereprepared:

-   -   α-(2-Ethoxyphenyl)-N-1-adamantylnitrone, and    -   α-(3-Ethoxy-4-methoxyphenyl)-N-1-adamantylnitrone.

Example 39 Synthesis of α-(4Ethoxyphenyl)-N-cyclopentylnitrone

4-Ethoxybenzaldehyde (22.0 g, 0.1467 mol) and N-cyclopentylhydroxylamine(14.1 g, 0.1398 mol) were mixed into toluene (200 mL) withp-toluenesulfonic acid monohydrate (1.0 g, 5.26 mmol). The mixture wasrefluxed for 3 hrs under argon atmosphere with a Dean-Stark trap toremove generated water. The solution was rotary evaporated to give aresidue which was purified by flash chromatography over silica gel withEtOAc as an eluant and then recrystallization from a mixed solvent ofhexanes and EtOAc. The title compound was obtained as a solid (21.24 g65.1% yield), m.p. 95.1° C. (R_(f)=0.18 on a silica gel plate usinghexanes:EtOAc, 2:1, v/v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2977 (CH), 2873 (CH), 1601 (C═N & benzene ring), 1575(benzene ring), 1251 (Ar—O) and 1169 (N—O).

¹H NMR (CDCl₃, 270 MHZ): δ=8.22 (2H, d, J=9.0 Hz, aromatic 2H), 7.41(1H, s, CH═N), 6.91 (2H,d, J=9.0 Hz, aromatic 2H), 4.40 (1H, tt, J=6.3 &7.8 Hz, CH), 4.07 (2H, q, J=7.0 Hz, OCH₂), 2.33-2.20 (2H, m, cyclopentyl2H), 2.04-1.86-(4H, m, cyclopentyl 4H), 1.70-154 (2H, m, cyclopentyl2H), and 1.42 (3H, t, J=7.0 Hz, CH₃) ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=160.13, 132.45, 130.39, 125.56, 114.19,75.72, 63.45, 31.25, 25.56 and 14.66 ppm.

Example 40 Synthesis of α-(4-Ethoxyphenyl)-N-tert-octylnitrone

The title compound was prepared by oxidation ofN-(4-ethoxyphenyl)-N-tert-octylamine with m-chloroperoxybenzoic acid inmethylene chloride. The amine was synthesized via NaBH₄ reduction fromthe corresponding imine which was acquired by condensation of4-ethoxybenzaldehyde and tert-octylamine in methanol. The title compoundwas isolated in 65.0% overall yield as white crystals, m.p. 100.8° C.(R_(f)=0.33 on silica gel plate using hexanes:EtOAc, 7:3, v/v, as aneluant).

Spectroscopic data were as follows:

IR (Kbr, cm⁻¹): 2978 & 2951 (CH), 1605 (C═N & benzene ring), 1563(benzene ring), 1263 (Ar—O) and 1114 (N—O).

¹H NMR (CDCl₃, 270 MHZ): δ=8.27 (2H, d, J=9.0 Hz, aromatic 2H), 7.49(1H, s, CH═N), 6.91 (2H,d, J=9.0 Hz, aromatic 2H), 4.08 (2H, q, J=7.0Hz, OCH₃), 1.97 (2H, s, CH₂), 1.64 (6H, s, 2 CH₃), 1.42 (3H, t, J=7.0Hz, CH₃) and 0.97, (9H, s, 3 CH₃) ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=160.12, 130.55, 130.17, 124.11, 114.20,73.20, 63.49, 51.35, 31.61, 30.69, 28.82 and 14.72 ppm.

Example 41 Synthesis of α-(4Benzyloxyphenyl)-N-tert-butylnitrone

A mixture of 4-benzyloxybenzaldehyde, N-tert-butylhydroxylamine andcatalytic amount of p-toluenesulfonic acid monohydrate in benzene wasrefluxed under argon atmosphere with a Dean-Stark trap to removegenerated water. The mixture was then rotary evaporated to give aresidue which was purified by recrystallization. The title compound wasobtained in 89.3% yield as a white powder, m.p. 111.0° C. (R_(f)=0.66 ona silica gel plate using EtOAc as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2982 (CH), 1601 (C═N & benzene ring), 1508 (benzenering), 1242 (Ar—O), 1170 (N—O). and 1005 (benzyl-O)

¹H NMR (CDCl₃, 270 MHZ): δ=8.29 (2H, d, J=9.2 Hz, aromatic 2H), 7.46(1H, s, CH═N), 7.41-7.31 (5H, m, aromatic 5H), 7.00 (2H, d, J=9.2 Hz,aromatic 2H),5.10 (2H, s, OCH₂ ), and 1.60 (9H, s, 3 CH₃)ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=159.89, 136.47, 130.64, 129.35, 128.52,128.00, 127.42, 124.23, 114.58, 70.05, 69.91 and 28.25 ppm

Example 42 Synthesis of α-(4-Benzyloxyphenyl)-N-cyclopentylnitrone

A mixture of 4-benzyloxybenzaldehyde (20 g, 94.23 mmol),N-cyclopentylhydroxylamine (14.3 g, 141.34 mmol), molecular sieves (60g, 4 A) and silica gel (15 g) in chloroform (300 mL) was stirred at roomtemperature under argon atmosphere for 48 hrs and then was refluxed foran additional 3 hrs. The mixture was filtered and rotary evaporated togive crystals which were recrystallized from hexanes and EtOAc toprovide the title compound as white crystals, 23.7 g, 85.1% yield), m.p.115.1° C. (R_(f)=0.35 on a silica gel plate using hexanes:EtOAc, 1:1,v:v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2953 (CH), 2867 (CH), 1601 (C═N & benzene ring), 1505(benzene ring), 1251 (Ar—O), 1139 (N—O). and 1009 (benzyl-O).

¹H NMR (CDCl₃, 270 MHZ): δ=8.23 (2H, d, J=9.2 Hz, aromatic 2H), 7.40(1H, s, CH═N), 7.43-7.27 (5H, m, aromatic 5H), 6.98 (2H, d, J=9.2 Hz,aromatic 2H), 5.08 (2H, s, OCH₂ ), 4.36 (1H, tt, J=7.7 & 6.1 Hz, CH),2.33-2.20 (2H, m, cyclopentyl 2H), 2.04-1.86 (4H, m, cyclopentyl 4H),1.70-1.54 (2H, m, cyclopentyl 2H) ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=159.79, 136.39, 132.21, 130.30, 128.47,127.94, 127.36, 123.96, 114.56, 75.71, 69.85, 31.21 and 25.50 ppm

Example 43 Synthesis of α-(4-Benzyloxyphenyl)-N-cyclohexylnitrone

The title compound was prepared according to the procedure described inExample 42 using 4-benzyloxybenzaldehyde and N-cyclohexylhydroxylamine.The title compound was obtained in 81.2% yield as slightly yellowishsolid, m.p. 129.0° C. (R_(f)=0.30 on a silica gel plate usinghexanes:EtOAc, 1:1, v:v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2993 (CH), 2854 (CH), 1603 (C═N & benzene ring), 1507(benzene ring), 1251 (Ar—O), 1138 (N—O). and 1012 (benzyl-O)

¹H NMR (CDCl₃, 270 MHZ): δ=8.24 (2H, d, J=8.9 Hz, aromatic 2H), 7.35(1H, s, CH═N), 7.44-7.32 (5H, m, aromatic 5H), 7.00.(2H, d, J=8.9 Hz,aromatic 2H), 5.11 (2H; s, OCH₂ ), 3.79 (1H, m, CH), 2.10-1.89 (6H, m,cyclopentyl 6H), 1.70 (1H, m, cyclopentyl 1H), and 1.22-1.45 (3H, m,cyclopentyl 3H) ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=159.90, 136.51, 131.70, 130.44, 128.58,128.05, 127.45, 124.05, 114.68, 75.14, 69.97, 31.12 and 25.09 ppm.

Example 44 Synthesis of α-(2-Ethoxyphenyl)-N-cyclopentylnitrone

The title compound was prepared according to the procedure described inExample 42 using 2-ethoxybenzaldehyde and N-cyclopentylhydroxylamine.The title compound was obtained in 72.6% yield as white crystals, m.p.87.3° C. (R_(f)=0.43 on a silica gel plate using-hexanes:EtOAc, 2:1,v:v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2976 (CH), 2957 (CH), 1636 (C═N), 1597 & 1564. (benzenering), 1251 (Ar—O), 1165 (N—O). and 1043 (Et—O).

¹H NMR (CDCl₃, 270 MHZ): δ=9.33 (1H, dd, J=7.8 & 1.7 Hz, aromatic 1H),7.98 (1H, s, CH═N), 7.32 (1H, ddd, J=8.2, 7.5 & 1.7 Hz, aromatic 1H),6.99 (1H, td, J=6.1 & 7.8 Hz, CH), 4.07 (2H, q, J=7.0 Hz, OCH₂ ),2.35-2.22 (2H, m, cyclopentyl 2H), 2.07-1.88 (4H, m, cyclopentyl 4H),1.72-1.57 (2H, m, cyclopentyl 2H) and 1.45 (3H, t, J=7.0 Hz, CH₃) ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=156.11, 131.00, 128.64, 127.31, 120.50,119.88, 110.57, 76.64, 63.85, 31.38, 25.55 and 14.74 ppm.

Example 45 Synthesis of α-(3-Ethoxy-4-methoxyphenyl)-N-tert-octylnitrone

A solution of 3-ethoxy-4methoxybenzaldehyde, N-tert-octylhydroxylamineand catalytic amount of HCl in methanol was refluxed for 90 hrs withmolecular sieves in a soxhlet for waster removal. The title compound wasobtained in 60.0% yield as white powder, m.p. 77.5° C. (R_(f)=0.40 on asilica gel plate using hexanes:EtOAc, 3:2, v:v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2975 (CH), 1636 (C═N), 1597 & 1575 (benzene ring), 1279(Ar—O), 1145.(N—O). and 1039 & 1026 (alkyl-O).

¹H NMR (CDCl₃, 270 MHZ): δ=8.44 (1H, d, J=2.0 Hz, aromatic 1H), 7.50(1H, dd, J=8.3 & 2.0 Hz, aromatic 1H), 7.48 (1H, s, CH═N), 6.90 (1H, d,J=8.3 HZ aromatic 1H), 4.20 (2H, q, J=7.OHz, OCH₂), 3.91 (3H, s, CH₃),1.97 (2H, s, CH₂), 1.64 (6H, s, 2CH₃), 1.48 (3H, t, J=7.0 Hz, CH₃), 0.98(9H, s, 3CH₃), ppm.

¹³C NMR (CDCl₃, 67.9 MHZ): δ=150.44, 147.71, 130.41, 124.56, 122.85,112.22, 110.67, 73.36, 64.23, 55.87, 51.43, 31.63, 30.68, 28.78 and14.72 ppm.

Example 46 Synthesis ofα-(3-Ethoxy-4-methoxyphenyl)-N-(2,4,-dimethyl-2-pentyl)nitrone

The title compound can be prepared according to the procedure describedin Example 45 using 3-ethoxy-4-methoxybenzaldehyde andN-2,4,-dimethyl-2-pentylhydroxylamine.

Example 47 Synthesis of α-[4-(4Fluorobenzyloxy)phenyl]-N-tert-butylnitrone

The title compound was prepared by refluxing a benzene solution of4-(4-fluorobenzyloxy)benzaldehyde and N-tert-butylhydroxylamine for 21hours with p-toluenesulfonic acid as a catalyst. The title compound wasobtained as a solid in 98.5% yield, m.p. 180.3° C. (R_(f)=0.16 on asilica gel plate using hexanes: EtOAc, 1:1, v/v, as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2984 (CH), 1607 (C═N & benzene ring), 1509 (benzenering), 1218 (Ar—O) and 1121 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.29 (2;H d, J=9.0 Hz, aromatic 2H), 7.47(1H, s, CH═N), 7.40 (2H, dd, J=8.7 & 5.3 Hz, aromatic 2H), 7.07 (2H, t,J=8.7 Hz, aromatic 2H), 6.99 (2H, d, J=9.0 Hz, aromatic 2H), 5.07 (2H,s, CH₂O) and 1.61 (9H, s, C(CH₃)₃) ppm.

¹³C NMR (CDCl₃, 67.9 MHz): δ=162.49 (d, J_(F)=246.5 Hz), 159.69, 132.26,130.66, 129.37, 129.27, 124.39, 115.47 (d, J_(F)=21.3 Hz), 114.55,70.12, 69.25, 28.27 ppm.

Example 48 Synthesis of α-(3-Ethoxy-4-methoxyphenyl)-N-cyclobutylnitrone

A solution of 3-ethoxy-4-methoxybenzaldehyde, cyclobutylaminehydrochloride salt, molecular sieves and silica gel in chloroform wasrefluxed for 20 hours. Filtration and rotary evaporation gave thecorresponding imine intermediate which was reduced with NaBH in ethanolto give N-cyclobutyl-N-(3-ethoxy-4-methoxybenzyl)amine. This amineintermediate was oxidized with H₂O₂/Na₂WO₄ in acetone/water to affordthe nitrone product. The title compound was obtained in 19.9% overallyield as cream-colored crystals, m.p. 112.7° C. (R_(f)=0.30 on a silicagel plate using EtOAc as an eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2980 (CH), 2935 (CH), 1634 (C═N), 1597 and 1586 (benzenering), 1265 (Ar—O), 1134 (N—O) and 1047 and 1021 (alkyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=8.48 (1H, d, J=2.0 Hz, aromatic 1H), 7.42(1H, dd, J=8.5 and 2.0 Hz, aromatic 1H), 7.30 (1H, s, CH═N), 6.89 (1H,d, J=8.5 Hz, aromatic 1H), 4.53 (1H, quintet, J=8.1 Hz, cyclobutyl CH),4.19 (2H, q, J=7.0 Hz, OCH₂) 3.91 (3H, s, CH₃), 2.84-2.68 (2H, m,cyclobutyl 2H), 2.36-2.25 (2H, m, cyclobutyl 2H), 1.91-1.75 (2H, m,cyclobutyl 2H), 1.48 (3H, t, J=7.0, CH₃) ppm.

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.68, 147.73, 132.30, 123.77, 122.86,112.11, 110.71, 67.45, 64.18, 55.83, 26.97, 14.69 and 14.15 ppm.

Example 49 Synthesis of α-(3-Ethoxy-4-methoxyphenyl3-N-(4-methylpent-2-yl)nitrone

3-Ethoxy-4-methoxybenzaldehyde (12.0 g, 0.0666 mol) andN-(4-methylpent-2-yl)hydroxylamine (9.36 g, 0.0799 mol) were mixed intobenzene (200 mL) with p-toluenesulfonic acid monohydrate (1.0 g, 5.26mol.). The mixture was refluxed for 16 hours under an argon atmospherewith a Dean-Stark trap to remove the generated water. The solution wasrotary evaporated, dissolved in ethyl acetate, washed with 5% aqueoussodium hydroxide solution, dried over magnesium sulfate, filtered andconcentrated. The title compound was obtained as a white solid (17.07 g,91.8% yield), m.p. 87.2° C. (R_(f)=0.31 on a silica gel plate usinghexanes;EtOAc, 1: 1, v/v, as an eluant). TheN-(4-methyl-2-pentyl)hydroxylamine precursor was obtained by sodiumcyanoborohydride reduction of 4-methyl-2-pentanone oxime in methanol,with hydrochloric acid catalysis.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2962 (CH), 1632 (C═N and benzene ring). 1588 (benzenering), 1265 (Ar—O) and 1129 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.44 (1H, d, J=2.0 HZ, aromatic H), 7.45 (1H,dd, J=8.5 and 2.0 Hz, aromatic H), 7.33 (1H, s, CH═N), 6.89 (H, d, J=8.5Hz, aromatic H), 4.19 (2H, q, J=7.0 Hz, OCH₂), 4.07 (1H, m, N(O)CH),3.91 (3H, s, OCH₃), 2.09-1.99 (1H, m, pentyl C⁴H), 1.66-1.34 (8H, m, CH₃of EtO, pentyl C¹H₃, and pentyl C³H₂), 0.95 (3H, d, J=8.4 Hz, pentylC⁵H₃), and 0.94 (3H, d, J=8.6 Hz, 4-methyl of pentyl) ppm.

¹³C NMR (CDCl₃, 67.9 MHz): δ=150.55, 117.75, 132.59, 123.88, 122.64,112.12, 110.69, 69.93, 64.19, 55.83, 43.09, 24.74, 22.80, 22.10, 19.64,14.67 ppm.

Example 50 Synthesis of α-(4-Benzyloxyphenyl)-N-cyclooctylnitrone

A solution of 4-benzyloxybenzaldehyde (12.7 g, 0.060 mol),N-cyclooctylhydroxylamine (10.0 g 0.070 mol) and a catalytic amount ofHCl in methanol. (300 mL) was refluxed for 56 hours with molecularsieves in a soxhlet for water removal. The reaction mixture wasconcentrated and dry flash columned on silica with hexanes/ethyl acetateto give the title compound as a pale yellow powder, (9.53 g, 47.0%yield), m.p. 107.5° C. (R_(f)=0.46 on a silica gel plate usinghexanes:EtOAc, 1:1, v:v, as an eluant). The N-cyclooctylhydroxylamineprecursor was obtained by sodium cyanoborohydride reduction ofcyclooctanone oxime in acetic acid/tetrahydrofuran.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 3061 (CH), 2967 (CH), 1648 (C═N), 1.603 (benzene ring),1579 (benzene ring), 1251 (Ar—O) and 1147 (N—O).

1H NMR (CDCl₃, 67.9 MHz): δ=8.22 (2H, d, J=9.0 Hz, aromatic 2H),7.47-7.29 (6H, m, aromatic 5H & CH═N), 6.99 (2H, d, J=9.0 Hz, aromatic2H), 5.10 (2H, s, benzyl CH₂), 4.08-3.97 (1H, m, N(O)CH), 2.31-2.15 (2H,m, cyclooctyl), 2.10-1.97 (2H, m, cyclooctyl), 1.94-1.76 (2H, m,cyclooctyl), 1.76-1.40 (8H, m, cyclooctyl) ppm.

¹³C NMR (CDCl₃, 67.9 MHz): δ=159.80, 136.47, 131.27, 130.34, 128.56,128.02, 127.44, 124.07, 114.64, 76.73, 69.93, 31.96, 26.54, 26.01, 24.70ppm.

Examples 51-80

Using the procedures described herein and the appropriate startingmaterials, the following additional compounds were prepared:

-   α-(2-ethoxyphenyl)-N-benzylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-(2,2,4,4-tetramethylpent-3-yl)nitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-but-2-ylnitrone-   α-(2-ethoxyphenyl)-N-but-2-ylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-cyclopentylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-n-propylnitrone-   α-(4-benzyloxyphenyl)-N-n-propylnitrone-   α-(4-benzyloxyphenyl)-N-isopropylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-(2-methylbut-2-yl)nitrone-   α-(2-ethoxyphenyl)-N-(2-methylbut-2-yl)nitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-cyclooctylnitrone-   α-(2-ethoxyphenyl)-N-cyclobutylnitrone-   α-(4-benzyloxyphenyl)-N-cyclobutylnitrone-   α-(4-benzyloxyphenyl)-N-tert-octylnitrone-   α-[4-(4-fluorobenzyloxy)phenyl]-N-cyclohexylnitrone-   α-(2-ethoxyphenyl)-N-tert-octylnitrone-   α-[4-(4-fluorobenzyloxy)phenyl]-N-isopropylnitrone-   α-(2-ethoxyphenyl)-N-cyclooctylnitrone-   α-(4-benzyloxyphenyl)-N-cyclopropylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-cyclopropylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-(3,5-dimethyl-1-adamantyl)nitrone-   α-(4-benzyloxyphenyl)-N-1-adamantylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-(1-methoxy-2-methylprop-2-yl)nitrone-   α-(4-benzyloxyphenyl)-N-2-adamantylnitrone-   α-(4-ethoxyphenyl)-N-cyclooctylnitrone-   α-(4-ethoxyphenyl)-N-1-adamantylnitrone-   α-[4-(4-methoxybenzyloxy)phenyl]-N-tert-butylnitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-(3-methylbut-1-yl)nitrone-   α-(3-ethoxy-4-methoxyphenyl)-N-cyclooctylnitrone, and-   α-[4-(4-fluorobenzyloxy)phenyl]-N-cyclopentylnitrone.

Comparative Example 1 Synthesis ofα-(2-Methoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 7 using 2-methoxybenzaldehyde and N-tert-butylhydroxylamine. Thetitle compound was isolated in 82.9% overall yield as white crystals,m.p. 109.1° C. (R_(f)=0.53 on a silica gel plate using ethyl acetate asthe eluant).

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 3004.0 (aromatic CH), 2966.0 (CH), 1593.4 (C═N), 1556.1(benzene ring), 1235.4 (Ar—O), 1125.5 (N—O), and 1017.3 (alkyl-O).

¹H NMR (CDCl₃, 270 MHz): δ=9.343 (1H, dd, J=7.9 Hz, J=1.7. Hz, aromaticH), 8.025 (1H, s, nitronyl CH), 7.329 (1H, td, J=7.9 Hz, J=1.7 Hz,aromatic H), 6.993 (1H, t, J=7.7 Hz, aromatic H), 6.856 (1H, d, J=8.4Hz, aromatic H), 3.841 (3H, s, OCH₃) and 1.587 (9H, s, 3 CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=157.452, 131.353, 128.836, 124.535,120.890, 120.234, 109.739, 70.843, 55.392 and 28.058.

Comparative Example 2 Synthesis ofα-(3-Methoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 11 using 3-methoxybenzaldehyde and N-tert-butylhydroxylamine.The title compound was isolated in 56.5% overall yield as a crystallinesolid, m.p. 93.4° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2977 (CH), 1589 (C═N), 1110 (N—O) and 1035 (C—O).

¹H NMR (DMSO-d₆, 270 MHz): δ=8.20 (1H, m, phenyl H), 7.84 (1H, s,nitronyl H), 7.78 (1H, d, J=8.0 Hz, phenyl H), 7.33 (1H, t, J=8.0 Hz,phenyl H), 6.98 (1H, dd, J=8.1, 2.5 Hz, phenyl H), 3.77 (3H, s, CH₃) and1.51 (9H, s, 3 CH₃).

¹³C NMR (DMSO-d₆, 67.9 MHz): δ=159.76, 133.55, 129.80, 129.40, 121.96,116.41, 113.34, 70.89, 55.35 and 28.04.

Comparative Example 3 Synthesis of α-(4-Ethoxyphenyl)-N-isopropylnitrone

The title compound was prepared according to the procedure described inExample 11 using 4-ethoxybenzaldehyde and N-isopropylhydroxylamine. Thetitle compound was isolated in 41.2% yield as a solid, m.p. 115.1° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 2979.6 (CH), 1597.5 (C═N), 1302.4 (CH₃), 1259.2 (C—O—C)and 1169.4 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.20 (2H, d, J=9.0 Hz, phenyl 2CH), 7.33 (1H,s, nitronyl CH), 6.88 (2H, d, J=9.0 Hz, phenyl 2CH), 4.06 (3H, m, CH₂and CH), 1.46 (6H, m, 2 CH₃) and 1.40 (3H, m, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz): δ=160.6, 131.8, 130.7, 123.8, 114.4, 67.1,63.4, 20.5 and 14.3.

Comparative Example 4

Synthesis of α-(4-Butoxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 11 using 4-butoxybenzaldehyde and N-tert-butylhydroxylamine. Thetitle compound was isolated in 96% yield (7.18 g) as a solid, m.p. 68.5°C. Spectroscopic data were as follows:

¹H NMR (CDCl₃, 270 MHz): δ=8.27 (2H, d, J=8.8, phenyl 2H), 7.45 (1H, s,nitronyl H), 6.91 (2H, d, J=8.8 Hz, phenyl 2H), 4.00 (2 H, t, CH₂), 1.60(9H, s, tert-butyl H), 1.50 (4H, m, 2CH₂), 0.97 (3H, t, J=6.7 Hz, CH₃).

Comparative Example 5 Synthesis ofα-(4-Pentloxyphenyl)-N-tert-butylnitrone

The title compound was prepared according to the procedure described inExample 2 using 4-hydroxybenzaldehyde, 1-iodopentane and2-methyl-2-nitropropane. The title compound was isolated in 75% overallyield as a solid, m.p. 43.2° C.

Spectroscopic data were as follows:

IR (KBr, cm⁻¹): 3092.7 (CH), 2972.1 (CH), 1604.9 (C═N), 1362.9 (CH₃),1258.8 (C—O—C) and 1117.3 (N—O).

¹H NMR (CDCl₃, 270 MHz): δ=8.24 (2H, d, J=9.1 Hz, phenyl 2H), 7.43 (1H,s, nitronyl H), 6.69 (2H, s, J=9.1 Hz, phenyl 2H), 3.97 (2H, t, J=6.4Hz, CH₂), 1.76 (2H, m, CH₂), 1.57 (9H, s, 3 CH₃), 1.39 (4H, m, 2, CH₂)and 0.90 (3H, t, J=6.9 Hz, CH₃).

¹³C NMR (CDCl₃, 67.9 MHz) δ=160.8, 130.9, 129.7, 124.0, 114.4, 69.9,68.0, 28.5, 28.0, 27.8, 22.1 and 13.6.

Comparative Example 6 Synthesis ofα-(4-Hexyloxyphenyl)-N-tert-butylnitrone

A solution of 4-hexyloxybenzaldehyde (3.83 g, 18.6 mmol) in 120 mL ofbenzene was refluxed with N-tert-butylhydroxylamine (3.32 g, 37.2 mmol)for 18 hours. The reaction mixture was then concentrated by rotaryevaporation and the resulting residue was purified by silica gel columnchromatography using 50:50 ethyl acetate/hexane to afford the titlecompound (2.88 g, 55.8% yield) as a solid, m.p. 69.0° C.

Spectroscopic data were as follows:

¹H NMR (CDCl₃, 270 MHz) δ=8.27 (2H, d, J=8.8 Hz, phenyl 2H), 7.45 (1H,s, nitronyl H), 6.91 (2H, d, J=8.8 Hz, phenyl 2H), 4.00 (2H, t, J=6.4Hz, O—CH₂), 1.60 (9H, singlet, tert-butyl H), 1.36 (8H, m, 4 CH₂) and0.90 (3H, t, CH₃).

Example I Electron Spin Resonance (ESR) Study

In this experiment, the ability of α-aryl-N-alkylnitrones of formula Iabove to trap free radicals is demonstrated using ESR spin trappingtechniques. See, for example, K. R. Maples et al., “In Vivo Detection ofFree Radical Metabolites”, Free Radicals in Synthesis and Biology (F.Minisci, ed.) pp. 423-436 (Kluwer Academic Publishers, Boston, 1989);and J. A. DeGray et al., “Biological Spin Trapping”, Electron SpinResonance 14:246-300 (1994). A t-butyl hydroperoxide/ferrous iron freeradical generating system was used in this experiment. This free radicalgenerating system produces t-butyl-alkoxyl radicals, t-butyl-peroxylradicals, and methyl radicals. If the α-aryl-N-alkylnitrones of thisinvention are capable of trapping any of these radicals to form a stableradical adduct, such radical adducts should be detectable by ESRspectroscopy.

To 490 μl of a 100 mM solution of α-(2-ethoxyphenyl)-N-tert-butylnitronein water was added 5 μl of 100 mM ferrous sulfate. The reaction wasinitiated by the addition of 5 μl of 100 mM t-butyl hydroperoxide. Thefinal concentrations of reagents are 1 mM ferrous iron, 1 mM t-butylhydroperoxide and 98 mM of the nitrone compound in water. Once mixed,the solution was quickly transferred into a quartz flat cell and thiscell was placed in the cavity of a Bruker ESP 300 ESR spectrometer, andscanned within 5 minutes of mixing. ESR spectrometer settings were: 3480G center field, 200 G field width, 480 seconds sweep time, 9.76 GHzfrequency, 10 dB power, 1.6×10⁵ receiver gain, 0.200 G modulationamplitude, 0.320 second time constant, and 270° phase. The resulting ESRspectrum, as shown in FIG. 1, consisted of primarily one species,characterized as a 16.8 G (1:1:1) triplet of 4.3 G (1:1) doublets,representing a_(N) and a_(H), respectively. This species is believed tobe the methyl radical adduct of α-(2-ethoxyphenyl)-N-tert-butylnitrone.Thus, the ESR spectrum shown in FIG. 1 demonstrates that theα-aryl-N-alkylnitrones of formula I are effective at trapping freeradicals and that such compounds can be used as analytical reagents forESR applications.

Example II Inhibition of Aβ Beta-Pleated Sheet Formation

The deposition of amyloid β-peptide (Aβ) is associated with thedevelopment of Alzheimer's disease. See, for example, G. G. Glenner etal. (1984) Biochem. Biophys. Res. Commun., 120:885-890; and R. E. Tanzi(1989) Ann. Med., 21:91-94. Accordingly, compounds which effectivelydisrupt the formation of Aβ(140) or Aβ(142) beta-pleated sheets arepotentially useful for preventing and/or reversing such amyloiddeposits. Thioflavin T (ThT) is known to rapidly associate withbeta-pleated sheets, particularly the aggregated fibrils of syntheticAβ(142). This-association gives rise to an excitation maximum at 440 nmand to an emission at 490 nm. In this experiment, the ability of certainα-aryl-N-alkylnitrones of formula I above to inhibit the association ofThT with synthetic Aβ(142) is demonstrated by measuring changes influorescence.

The experiments were performed using a CytoFluor II fluorescence platereader having the following parameters: Filters: Excitation 440 nm/20Emission 490 nm/40 Gain: 75 Cycle to Cycle Time: 30 min Run Time: 720min (24 cycles) or dependent on experimental design Plate: 96 well

Into each well was aliquoted 95 μl of ThT (3 μM) prepared in PBS (pH6.0), 2 μL of the compound to be tested (10 μM) prepared with 0.05% ofmethylcellulose in PBS (pH 6.0), and 3 μL of Aβ(142)(3 μg) prepared withdH₂O. The fluorescence measurement began when the Aβ(142) was added andcontinued for a total of 12 hours. The percent inhibition ofbeta-pleated sheet formation was calculated from the relativefluorescence unit difference between aggregation in the presence and inthe absence of the test compounds. Inhibition of Aβ(1-42) beta-pleatedsheet formation by at least 30% compared to the controls is consideredsignificant in this test. The results of these in vitro tests aredescribed below.

Example III Protection Against Aβ(25-35) Induced Neuronal Cell Loss

Patients with Alzheimer's disease are known to suffer a progressive lossof neuronal cells. See, for example, P. J. Whitehause et al., (1982)Science, 215:1237-1239. In this experiment, the ability of certainα-aryl-N-alkylnitrones of formula I above to protect againstAβ(25-35)-induced neuronal cell loss is demonstrated. Sprague Dawley rathippocampus of 18-day-gestation embryos was excised and then dissociatedby trituration to prepare primary neuronal cultures. Cells (3×10⁵) wereplated on 35 mm poly-D-lysine-coated plates containing Eagle's minimumessential medium supplemented with 10% fetal bovine serum. After 3-5hours, the original medium was removed and replaced with 1 mL of freshmedium. Cultures were-maintained at 37° C. in a 5% CP₂/95% airhumidified incubator. Glial growth is observed as a monolayer underneurons.

To the cells (7 DIV) was added 30 μM of Aβ(25-35) dissolved in H₂O(stored at −20° C.) and 100 μM of the test compound in 1%methylcellulose. Controls were also conducted without the test compound.The percentage of morphologically viable neurons was determined bycounting the number of viable neurons after 96 hours treatment (threeregions/well, n=6 wells). Inhibition of Aβ(25-35)-induced neuronal cellloss by at least 30% compared to the controls is considered significantin this test. The results of these in vitro tests are described below.

Example IV Reduction of β-Amyloid-Induced Increased Release ofInterleukin-1β and Tumor Necrosis Factor-α

In this experiment, the ability of certain α-aryl-N-alkylnitrones offormula I above to reduce the β-amyloid-induced increased release overLPS alone of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNFα)is demonstrated. THP-1 cells, a human monocyte cell line from AmericanType Culture Collection, were grown in RPMI-1640 medium plus 10% fetalbovine serum (FBS, not heat-inactivated) in T-flasks. The medium waschanged every two days by spinning down the cells (800 rpm, 5 minutes)and adding the same fresh medium. Alternatively, the cultures weremaintained by supplementation with fresh medium. The cultures weremaintained at a cell concentration ranging from between 1×10⁵ and 1×10⁶cells/mL. Because sera may contain unknown factors which can affectmacrophage/monocyte IL-1 production, the FBS was reduced to 5% for 24hours. The FBS was further reduced to 2% over two days prior to startingeach experiment. The cells were collected by centrifugation andresuspended in media containing 2% FBS. Cell numbers were calculated andcells were plated on 24-well plates (3×10⁵ cells/0.6 mL/well). Cellswere then treated with LPS (0.5 μg/ml or -0-10 μg/ml for LPSdose-response experiments) alone or in combination with Aβ peptides (5μM or 0.05-5 μM for dose-response experiments). When determining theeffect of the test compounds on IL-1β and TNFα release, 10 μM of thetest compound was added with the LPS and Aβ(25-35) and this mixture wasincubated for 48 hours prior to performing ELISA.

IL-1β and TNFα secretions into medium by LPS-stimulated THP-1 cells, inthe presence or absence of amyloid peptides and a test compound, wereassayed with a commercially available ELISA kit (R & D Systems).Briefly, a microtiter plate coated with a murine monoclonal antibody tohuman IL-1β or TNFα was supplied by the manufacturer. Standards andsamples were pipetted into the wells and any IL-1β or TNFα present wasbound by the immobilized antibody. Unbound proteins were washed away anda horseradish peroxidase-linked polyclonal antibody specific for IL-1βor TNFα was added to the wells to “sandwich” the IL-1β and TNFα bound inthe initial step. After washing to remove any unbound antibody-enzymereagent, a substrate solution (1:1 hydrogenperoxide:tetramethylbenzidine, v/v) was added to the wells and colordeveloped in proportion to the amount of IL-1β or TNFα bound in theinitial step. Color development was stopped with 2 N sulfuric acid andthe optical density of the standard and the test samples was measured at450 nm. The amounts of IL-1β or TNFα present in the samples werecalculated based upon a standard curve. Assays were run in quadruplicatewells. Inhibition of β-amyloid-induced increase release ofinterleukin-1β or tumor necrosis factor by at least 30% compared tocontrols is considered significant in these tests. The results of thesein vitro tests are described below.

Example V Protection Against IL-1β and IFNγ-Induced Toxicity

In this experiment, the ability of certain α-aryl-N-alkylnitrones offormula I above to reduce the IL-1β and IFNγ-induced neuronal toxicityin mixed rat hippocampal neuronal cultures is demonstrated. Rathippocampus of 18-day-gestation embryos were dissected free andincubated in HBSS containing 0.1% trypsin at 37° C. for 30 minutes.Tissue was then suspended in plating medium consisting of Eagle'sminimum essential medium supplemented with 2 mM L-glutamine, 14.75 mMKCl, 1 mM pyruvic acid, 10% fetal bovine serum and 100 units/mLpenicillin/100 μg/mL streptomycin. After trituration through aflame-polished Pasteur pipette, cells were diluted in additional platingmedium, counted and seeded at a density of 3.5×10⁵/mL/well on Falcon6-well plates which were precoated with 20 μg/mL poly-D-lysine for 2-3hours at room temperature, and washed twice with HBSS. After 3-5 hours,the original medium was removed and replaced with 1 mL of fresh medium.Cultures were maintained at 37° C. in a 5% CO₂/95% air humidifiedincubator for 12 days.

12 DIV hippocampal cultures which contain neurons and astrocytes wereused to perform the experiment. In each well was added 200 U/mL ofrecombinant mouse IL-1β (Genzyme) and 1,000 U/mL of IFNγ (Genzyme). 10μL of the test compound (100 μM final concentration) in 1% methylcellulose was added immediately to each well. To control wells wereadded only 1% methyl cellulose. Dexamethasone (30 μM) was used as apositive control. Cultures were incubated at 37° C. in a humidifiedatmosphere containing 5 % CO₂ for 48 or 96 hours. Neuronal injury wasestimated in all experiments by examination of cultures withphase-contrast-microscopy and was quantified by measurement of cytosoliclactate dehydrogenase (LDH) release into the culture medium.

Release of LDH into the bathing medium was estimated from the conversionof NAD to NADH, after lactate addition, and was measuredspectrophotometrically from the rate of decrease in 340 nm absorbance.LDH activity is defined as that amount of enzyme that catalyzed theformation of one micromole of NADH per minute under the conditions ofthe assay procedure. To a 96-well plate, 0.05 mL medium collected fromeach sample was added to and then mixed with 0.10 mL reagent from LD-L20 kit (Sigma). The plate was immediately placed into the SpectraMax 340plate reader to read at 340 nm wavelength at 25° C. for 3 minutes at 30second intervals. Reduction of neuronal injury by at least 30% comparedto controls is considered significant in this test. The results of thisin vitro test are described below.

In vitro Test Results:

Certain of the compounds prepared in the above examples were tested inat least one of the above described in vitro tests. The compounds ofthis invention either inhibited Aβ(1-42) beta-pleated sheet formationand/or Aβ(25-35)-induced neuronal cell loss and/or β-amyloid-inducedincrease release of interleukin-1β and/or tumor necrosis factor and/orIL-1β/IFN_(γ)-induced toxicity by at least 30% compared to the controlsor are expected to be effective in at least one of these in vitro assaysupon further testing. In contrast, the compounds of Comparative Examples1-6 failed to inhibit Aβ(1-42) beta-pleated sheet formation and/orAβ(25-35)-induced neuronal cell loss and/or β-amyloid-induced increaserelease of interleukin-1β and/or tumor necrosis factor and/orIL-1β/IFN_(γ)-induced toxicity by at least 30% compared to the controls.

Example VI Reduction of Cognitive Defects Due to Aβ-Peptide/Ibotenate

In this experiment, the ability of certain α-aryl-N-alkylnitrones offormula I above to reduce the in vivo impairment of animals treated withibotenate and Aβ(25-35) is demonstrated. The procedures employed in thisexample are similar to those described in Dornan et al., NeuroReport 5,165-168 (1993). Male Sprague-Dawley rats (200-300 g) were weighed andgiven 10 mg/kg of α-(2-ethoxyphenyl)-N-tert-butylnitrone or 1%methylcellulose by oral gavage. One hour later, the rats werestereotaxically injected into the CA1 region of their hippocampus with 8nmol of Aβ(25-35) and 6 nmol of ibotenate per side (coordinates frombregma −3.6=AP, ±2.2=ML, −3.0=DV from the top of the dura). Controlswere injected with PBS (pH 7.4). All injections were 1.5 μL in volume.The animals receiving. PBS were orally dosed with 1% methylcellulose.Oral dosing continued daily until the end of the Morris water mazetesting.

Nine to eleven days following injection, animals were tested in a Morriswater maze task to measure spatial memory and learning. Animals weregiven three days of testing with four to six trials per day. The lasttrial on the fourth day was a probe trial where the platform was removedand time in quadrant and annulae crossings were determined. Followingthe behavioral testing, animals were perfused with 10% neutral formalin.The brain was post-fixed for 1 week in 10% formalin and then sliced forhistological evaluation. Image analysis of cresyl violet staining wasused to compare the neuronal loss (lesion volume) in the hippocampusbetween groups. The data show thatα-(2-ethoxyphenyl)-N-tert-butylnitrone reduced the Aβpeptide/ibotenate-induced learning deficit.

Example VII Reduction of Cognitive Deficits in Autoimmune Mice

In this experiment, the ability of certain α-aryl-N-alkylnitrones offormula I above to reduce cognitive deficits in autoimmune strainsof-mice is demonstrated. MRL/MpJfas^(lpr) (“mutant mice” or “Fas^(lpr)”)strains of mice have been described as useful models of Lupus due totheir autoimmune lymphoproliferative pathology. In particular, themutant mice show a cognitive deficit at approximately four months ofage, which is not observed at two months of age. See, for example,Forster et al., 1988, Behav. Neural Biology, 49, 139-151.

In the experiment, male MRL/MpJ Fas^(lpr) and normal MRL/MpJ++ mice of 8weeks of age were weighed and administered 100 mg/kg of the testcompound (either α-(2-ethoxyphenyl)-N-tert-butylnitrone orα-(4-ethoxyphenyl)-N-cyclohexylnitrone) or 1% methylcellulose vehicle byoral gavage daily for 8 to 9 weeks. At 4 months of age, the mice weretested for avoidance, discrimination, session criteria and acquisitionin a one day-T-maze task with a maximum of 25 trials. Criteria was metwith four of five trials correct with the last two correct trials beingconsecutive in avoidance and discrimination. The Fas^(lpr) mice showed adeficit in both avoidance and acquisition compared to the normal micewhich received the 1% methylcellulose. In contrast, the Fag^(lpr) micetreated with the test compounds of this invention had reducedacquisition values and acquired avoidance skills earlier than untreatedmutant mice (i.e., similar to the normal controls). These resultsdemonstrate that α-aryl-N-alkylnitrones of formula I above reduced thecognitive deficits of the autoimmune strains of mice.

Example VIII Prevention of MBP-Induced Experimental AllergicEncephalomyelitis

Multiple sclerosis (MS) is a chronic inflammatory CNS disorder caused bydemyelination in the brain and spinal cord. The disease is characterizedby progressive CNS dysfunction, including muscular weakness, tremor,incontinence, ocular disturbances, and mental dysfunction, withremissions and exacerbations.

Experimental allergic encephalomyelitis (EAE) induced by injection ofguinea pig myelin basic protein (MBP) or MBP peptide fragments isreported to be a useful model for MS. See, for example, D. E. McFarlinet al., “Recurrent Experimental Allergic Encephalomyelitis in the LewisRat,” The Journal of Immunology, 113(2): 712-715 (1974). In thisexperiment, the ability of certain α-aryl-N-alkylnitrones of formula Iabove to prevent MBP-induced EAE is demonstrated.

Female, Lewis rats of 8 weeks of age (180-250 g) were weighed and thengiven two intradermal injections (0.1 mL each) of 0.4 mg of M.tuberculin in 0.1 mL incomplete Freunds adjuvant and 50 mg of myelinbasic protein in 0.1 mL of saline into the base of the tail. Animalswere weighed daily and given a clinical score beginning on Day 8, postinoculation, according to the following criteria:

-   -   0.0=No illness    -   0.5=Tip of tail flaccid    -   1.0=Entire tail flaccid    -   1.5=Hind limb weakness    -   2.0=Hind limb paralysis    -   2.5=Hind limb paralysis and front limb-weakness    -   3.0=Hind and front limb paralysis    -   4.0=Moribund state or death

On day 3, post-inoculation animals were administered b.i.d either a testcompound (100 mg/kg) or 1% methylcellulose vehicle by oral gavage up toand including day 16. The results demonstrate that the compounds ofExamples 3, 11, 17, 22, 41, 42 and 45 reduced the CNS inflammatorydeficit in acute EAE animals. At the dosages tested, the compounds ofExamples 4, 5, 7, 10, 15 and 29 did not significantly reduce the CNSinflammatory deficit.

From the foregoing description, various modifications and changes in thecompositions and methods of this invention will occur to those skilledin the art. All such modifications coming within the scope of theappended claims are intended to be included therein.

1-44. (canceled)
 45. A method for ameliorating a cause of aneurodegenerative disease in a patient at risk for developing theneurodegenerative disease which method comprises administering to saidpatient a pharmaceutical composition comprising a pharmaceuticallyacceptable carrier and an effective neurodegenerativedisease-cause-ameliorating amount of a compound of formula I:

wherein R¹ is selected from the group consisting of alkoxy, alkaryloxy,alkcycloalkoxy, aryloxy, and cycloalkoxy; R² is selected from the groupconsisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxy and halogen,or when R¹ and R² are attached to adjacent carbon atoms, R¹ and R² maybe joined together to form an alkylenedioxy group; R³ is selected fromthe group consisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxyand halogen; R⁴ is selected from the group consisting of hydrogen andalkyl; R⁵is selected from the group consisting of alkyl having at least3 carbon atoms, substituted alkyl having at least 3 carbon atoms andcycloalkyl; provided that: (i) when R² and R³ are independently hydrogenor methoxy, R¹ is not methoxy; (ii) when R², R³ and R⁴ are hydrogen andR⁵ is tert-butyl, then R¹ is not 4-n-butoxy, 4-n-pentyloxy or4-n-hexyloxy; (iii) when R², R³ and R⁴ are hydrogen and R⁵is isopropyl,then R¹ is not 4-ethoxy; (iv) when R¹ and R² are joined together to forma 3,4-methylenedioxy group and R³ are R⁴ are hydrogen, then R⁵ is notisopropyl or tert-butyl; (v) when R², R³ and R⁴ are hydrogen and R⁵ is1-hydroxy-2-methylprop-2-yl, then R¹ is not 2-ethoxy; (vi) when R¹ is4-methoxy, R² is 3-ethoxy, and R³ and R⁴ are hydrogen, then R⁵ is not2,2-dimethylbut-3-yl or 1-hydroxy-2-methylprop-2-yl; and (vii) when R³and R⁴ are hydrogen and R⁵ is tert-butyl, then R¹ is not 4-methoxy whenR² is 2-fluoro, and R¹ is not 2-methoxy when R² is 4-fluoro.
 46. Themethod according to claim 45, wherein the neurodegenerative disease isAlzheimer's disease.
 47. The method according to claim 45, wherein theneurodegenerative disease is Parkinson's disease.
 48. The methodaccording to claim 45, wherein the neurodegenerative disease is HIVdementia.
 49. (canceled)
 50. A method for ameliorating a cause of anautoimmune disease in a patient at risk for developing the autoimmunedisease which method comprises administering to said patient apharmaceutical composition comprising a pharmaceutically acceptablecarrier and an effective autoimmune disease-cause-ameliorating amount ofa compound of formula I:

wherein R¹ is selected from the group consisting of alkoxy, alkaryloxy,alkcycloalkoxy, aryloxy, and cycloalkoxy; R² is selected from the groupconsisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxy and halogen,or when R¹ and R² are attached to adjacent carbon atoms, R¹ and R² maybe joined together to form an alkylenedioxy group; R³ is selected fromthe group consisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxyand halogen; R⁴ is selected from the group consisting of hydrogen andalkyl; R⁵ is selected from the group consisting of alkyl having at least3 carbon atoms, substituted alkyl having at least 3 carbon atoms andcycloalkyl; provided that: (i) when R² and R³ are independently hydrogenor methoxy, R¹ is not methoxy; (ii) when R², R³ and R⁴ are hydrogen andR⁵ is tert-butyl, then R¹ is not 4-n-butoxy, 4-n-pentyloxy or4-n-hexyloxy; (iii) when R², R³ and R⁴ are hydrogen and R⁵ is isopropyl,then R¹ is not 4-ethoxy; (iv) when R¹ and R² are joined together to forma 3,4-methylenedioxy group and R³ are R⁴ are hydrogen, then R⁵ is notisopropyl or tert-butyl; (v) when R², R³ and R⁴ are hydrogen and R⁵ is1-hydroxy-2-methylprop-2-yl, then R¹ is not 2-ethoxy; (vi) when R¹ is4-methoxy, R² is 3-ethoxy, and R³ and R⁴ are hydrogen, then R⁵ is not2,2-dimethylbut-3-yl or 1-hydroxy-2-methylprop-2-yl; and (vii) when R³and R⁴ are hydrogen and R⁵ is tert-butyl, then R¹ is not 4-methoxy whenR² is 2-fluoro, and R¹ is not 2-methoxy when R² is 4-fluoro.
 51. Themethod according to claim 50, wherein the autoimmune disease is systemiclupus.
 52. The method according to claim 50, wherein the autoimmunedisease is multiple sclerosis.
 53. (canceled)
 54. A method forameliorating a cause of an inflammatory disease in a patient at risk fordeveloping the inflammatory disease which method comprises administeringto said patient a pharmaceutical composition comprising apharmaceutically acceptable carrier and an effective inflammatorydisease-cause-ameliorating amount of a compound of formula I:

wherein R¹ is selected from the group consisting of alkoxy, alkaryloxy,alkcycloalkoxy, aryloxy, and cycloalkoxy; R² is selected from the groupconsisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxy and halogen,or when R¹ and R² are attached to adjacent carbon atoms, R¹ and R² maybe joined together to form an alkylenedioxy group; R³is selected fromthe group consisting of hydrogen, alkoxy, alkcycloalkoxy, cycloalkoxyand halogen; R⁴is selected from the group consisting of hydrogen andalkyl; R⁵is selected from the group consisting of alkyl having at least3 carbon atoms, substituted alkyl having at least 3 carbon atoms andcycloalkyl; provided that: (i) when R² and R³ are independently hydrogenor methoxy, R¹ is not methoxy; (ii) when R², R³ and R⁴ are hydrogen andR⁵is tert-butyl, then R¹ is not 4-n-butoxy, 4-n-pentyloxy or4-n-hexyloxy; (iii) when R², R³ and R⁴ are hydrogen and R⁵ is isopropyl,then R¹ is not 4-ethoxy; (iv) when R¹ and R² are joined together to forma 3,4-methylenedioxy group and R³ are R⁴ are hydrogen, then R⁵ is notisopropyl or tert-butyl; (v) when R², R³ and R⁴ are hydrogen and R⁵ is1-hydroxy-2-methylprop-2-yl, then R¹ is not 2-ethoxy; (vi) when R¹ is4-methoxy, R² is 3-ethoxy, and R³ and R⁴ are hydrogen, then R⁵ is not2,2-dimethylbut-3-yl or 1-hydroxy-2-methylprop-2-yl; and (vii) when R³and R⁴ are hydrogen and R⁵ is tert-butyl, then R¹ is not 4-methoxy whenR² is 2-fluoro, and R¹ is not 2-methoxy when R² is 4-fluoro.
 55. Themethod according to claim 54, wherein the inflammatory disease isrheumatoid arthritis.
 56. The method according to claim 54, wherein theinflammatory disease is septic shock.
 57. The method according to claim54, wherein the inflammatory disease is nodosum leprosy.
 58. The methodaccording to claim 54, wherein the inflammatory disease is septicemia.59. The method according to claim 54, wherein the inflammatory diseaseis uveitis.
 60. The method according to claim 54, wherein theinflammatory disease is adult respiratory distress syndrome.
 61. Themethod according to claim 54, wherein the inflammatory disease isinflammatory bowel disease.